Pyrazole compounds as CB2 agonists

ABSTRACT

The invention relates to a compound of formula (I) 
                         
wherein A 1  to A 3  and R 1  to R 3  are defined as in the description and in the claims. The compound of formula (I) can be used as a medicament.

CROSS-REFERENCE TO RELATED APPLICATIONS

This application is a continuation application of InternationalApplication No. PCT/EP2014/054107, filed on Mar. 4, 2014, which claimspriority to European Patent Application No. 13158233.0, filed on Mar. 7,2013, the entire contents of which are incorporated herein by reference.

The present invention relates to organic compounds useful for therapyand/or prophylaxis in a mammal, and in particular to compounds that arepreferential agonists of the Cannabinoid Receptor 2.

The invention relates in particular to a compound of formula (I)

wherein

-   -   A¹ is carbon or nitrogen;    -   A² is carbon or nitrogen;    -   A³ is —(CH₂)_(n)— or —CH₂C(O)—;    -   R¹ is alkyl, cycloalkyl, alkoxy or halogen;    -   R² is alkoxy, substituted pyrrolidinyl or substituted        dihydropyrrolyl, wherein substituted pyrrolidinyl and        substituted dihydropyrrolyl are pyrrolidinyl and dihydropyrrolyl        substituted with one or two substituents independently selected        from halogen, hydroxyl, hydroxyalkyl, alkoxyalkyl and        alkylfurazanylalkoxy;    -   R³ is phenyl, substituted phenyl, substituted furazanyl,        pyridinyl, substituted pyridinyl, dioxothietanyl,        tetrahydrofuranyl, substituted tetrazolyl or substituted        triazolyl, wherein substituted phenyl, substituted furazanyl,        substituted pyridinyl and substituted triazolyl are phenyl,        pyridinyl and triazolyl substituted with one or two substituents        independently selected from alkyl, alkoxy, halogen, haloalkyl,        alkylsulfonyl and cycloalkyl, and wherein substituted tetrazolyl        and substituted furazanyl are tetrazolyl and furazanyl        substituted with one substituent selected from alkyl, alkoxy,        halogen, haloalkyl, alkylsulfonyl and cycloalkyl;    -   n is 0, 1 or 2;    -   provided that A¹ and A² are not both carbon at the same time;    -   or a pharmaceutically acceptable salt or ester thereof.

The compound of formula (I) is particularly useful in the treatment orprophylaxis of e.g. pain, atherosclerosis, age-related maculardegeneration, diabetic retinopathy, glaucoma, retinal vein occlusion,retinopathy of prematurity, ocular ischemic syndrome, geographicatrophy, diabetes mellitus, inflammation, inflammatory bowel disease,ischemia-reperfusion injury, acute liver failure, liver fibrosis, lungfibrosis, kidney fibrosis, systemic fibrosis, acute allograft rejection,chronic allograft nephropathy, diabetic nephropathy,glomerulonephropathy, cardiomyopathy, heart failure, myocardialischemia, myocardial infarction, systemic sclerosis, thermal injury,burning, hypertrophic scars, keloids, gingivitis pyrexia, livercirrhosis or tumors, regulation of bone mass, amyotrophic lateralsclerosis, multiple sclerosis, Alzheimer's disease, Parkinson's disease,stroke, transient ischemic attack or uveitis.

The compound of formula (I) is in particular useful in the treatment orprophylaxis of diabetic retinopathy, retinal vein occlusion or uveitis.

The cannabinoid receptors are a class of cell membrane receptorsbelonging to the G protein-coupled receptor superfamily. There arecurrently two known subtypes, termed Cannabinoid Receptor 1 (CB1) andCannabinoid Receptor 2 (CB2). The CB1 receptor is mainly expressed inthe central nervous (i.e. amygdala cerebellum, hippocampus) system andto a lesser amount in the periphery. CB2, which is encoded by the CNR2gene, is mostly expressed peripherally, on cells of the immune system,such as macrophages and T-cells (Ashton, J. C. et al. CurrNeuropharmacol 2007, 5(2), 73-80; Miller, A. M. et al. Br J Pharmacol2008, 153(2), 299-308; Centonze, D., et al. Curr Pharm Des 2008, 14(23),2370-42), and in the gastrointestinal system (Wright, K. L. et al. Br JPharmacol 2008, 153(2), 263-70). The CB2 receptor is also widelydistributed in the brain where it is found primarily on microglia andnot neurons (Cabral, G. A. et al. Br J Pharmacol 2008, 153(2): 240-51).

The interest in CB2 receptor agonists has been steadily on the riseduring the last decade (currently 30-40 patent applications/year) due tothe fact that several of the early compounds have been shown to havebeneficial effects in pre-clinical models for a number of human diseasesincluding chronic pain (Beltramo, M. Mini Rev Med Chem 2009, 9(1),11-25), atherosclerosis (Mach, F. et al. J Neuroendocrinol 2008, 20Suppl 1, 53-7), regulation of bone mass (Bab, I. et al. Br J Pharmacol2008, 153(2), 182-8), neuroinflammation (Cabral, G. A. et al. J LeukocBiol 2005, 78(6), 1192-7), ischemia/reperfusion injury (Pacher, P. etal. Br J Pharmacol 2008, 153(2), 252-62), systemic fibrosis(Akhmetshina, A. et al. Arthritis Rheum 2009, 60(4), 1129-36;Garcia-Gonzalez, E. et al. Rheumatology (Oxford) 2009, 48(9), 1050-6),liver fibrosis (Julien, B. et al. Gastroenterology 2005, 128(3), 742-55;Munoz-Luque, J. et al. J Pharmacol Exp Ther 2008, 324(2), 475-83).

Ischemia/reperfusion (I/R) injury is the principal cause of tissuedamage occurring in conditions such as stroke, myocardial infarction,cardiopulmonary bypass and other vascular surgeries, and organtransplantation, as well as a major mechanism of end-organ damagecomplicating the course of circulatory shock of various etiologies. Allthese conditions are characterized by a disruption of normal bloodsupply resulting in an insufficient tissue oxygenation. Re-oxygenatione.g., reperfusion is the ultimate treatment to restore normal tissueoxygenation. However the absence of oxygen and nutrients from bloodcreates a condition in which the restoration of circulation results infurther tissue damage. The damage of reperfusion injury is due in partto the inflammatory response of damaged tissues. White blood cells,carried to the area by the newly returning blood, release a host ofinflammatory factors such as interleukins as well as free radicals inresponse to tissue damage. The restored blood flow reintroduces oxygenwithin cells that damages cellular proteins, DNA, and the plasmamembrane.

Remote ischemic preconditioning (RIPC) represents a strategy forharnessing the body's endogenous protective capabilities against theinjury incurred by ischemia and reperfusion. It describes the intriguingphenomenon in which transient non-lethal ischemia and reperfusion of oneorgan or tissue confers resistance to a subsequent episode of “lethal”ischemia reperfusion injury in a remote organ or tissue. The actualmechanism through which transient ischemia and reperfusion of an organor tissue confers protection is currently unknown although severalhypotheses have been proposed.

The humoral hypothesis proposes that the endogenous substance (such asadenosine, bradykinin, opioids, CGRP, endocannabinoids, Angiotensin I orsome other as yet unidentified humoral factor) generated in the remoteorgan or tissue enters the blood stream and activates its respectivereceptor in the target tissue and thereby recruiting the variousintracellular pathways of cardioprotection implicated in ischemicpreconditioning.

Recent data indicates that endocannabinnoids and their receptors, inparticular CB2 might be involved in pre-conditioning and contribute toprevent reperfusion injury by downregulation of the inflammatoryresponse (Pacher, P. et al. Br J Pharmacol 2008, 153(2), 252-62).Specifically, recent studies using CB2 tool agonists demonstrated theefficacy of this concept for reducing the PR injury in the heart (Defer,N. et al. Faseb J 2009, 23(7), 2120-30), the brain (Zhang, M. et al. JCereb Blood Flow Metab 2007, 27(7), 1387-96), the liver (Batkai, S. etal. Faseb J 2007, 21(8), 1788-800) and the kidney (Feizi, A. et al. ExpToxicol Pathol 2008, 60(4-5), 405-10).

Moreover, over the last few years, a growing body of literatureindicates that CB2 can also be of interest in sub-chronic and chronicsetting. Specific upregulation of CB1 and CB2 has been shown to beassociated in animal models of chronic diseases associated with fibrosis(Garcia-Gonzalez, E. et al. Rheumatology (Oxford) 2009, 48(9), 1050-6;Yang, Y. Y. et al. Liver Int 2009, 29(5), 678-85) with a relevantexpression of CB2 in myofibroblasts, the cells responsible for fibrosisprogression.

Activation of CB2 receptor by selective CB2 agonist has in fact beenshown to exert anti-fibrotic effect in diffuse systemic sclerosis(Garcia-Gonzalez, E. et al. Rheumatology (Oxford) 2009, 48(9), 1050-6)and CB2 receptor has emerged as a critical target in experimental dermalfibrosis (Akhmetshina, A. et al. Arthritis Rheum 2009, 60(4), 1129-36)and in in liver pathophysiology, including fibrogenesis associated withchronic liver diseases (Lotersztajn, S. et al. Gastroenterol Clin Biol2007, 31(3), 255-8; Mallat, A. et al. Expert Opin Ther Targets 2007,11(3), 403-9; Lotersztajn, S. et al. Br J Pharmacol 2008, 153(2),286-9).

The compounds of the invention bind to and modulate the CB2 receptor andhave lower CB1 receptor activity.

In the present description the term “alkyl”, alone or in combination,signifies a straight-chain or branched-chain alkyl group with 1 to 8carbon atoms, particularly a straight or branched-chain alkyl group with1 to 6 carbon atoms and more particularly a straight or branched-chainalkyl group with 1 to 4 carbon atoms. Examples of straight-chain andbranched-chain C₁-C₈ alkyl groups are methyl, ethyl, propyl, isopropyl,butyl, isobutyl, tert.-butyl, the isomeric pentyls, the isomeric hexyls,the isomeric heptyls and the isomeric octyls, particularly methyl,ethyl, propyl, butyl and pentyl. Particular examples of alkyl are methyland tert.-butyl.

The term “cycloalkyl”, alone or in combination, signifies a cycloalkylring with 3 to 8 carbon atoms and particularly a cycloalkyl ring with 3to 6 carbon atoms. Examples of cycloalkyl are cyclopropyl, cyclobutyl,cyclopentyl and cyclohexyl, cycloheptyl and cyclooctyl. A particularexample of “cycloalkyl” is cyclopropyl.

The term “alkoxy”, alone or in combination, signifies a group of theformula alkyl-O— in which the term “alkyl” has the previously givensignificance, such as methoxy, ethoxy, n-propoxy, isopropoxy, n-butoxy,isobutoxy, sec-butoxy, tert.-butoxy and neopentyloxy. Particular“alkoxy” are methoxy, ethoxy and neopentyloxy.

The term “oxy”, alone or in combination, signifies the —O— group.

The terms “halogen” or “halo”, alone or in combination, signifiesfluorine, chlorine, bromine or iodine and particularly fluorine,chlorine or bromine, more particularly fluorine and chlorine. The term“halo”, in combination with another group, denotes the substitution ofsaid group with at least one halogen, particularly substituted with oneto five halogens, particularly one to three halogens, i.e. one, two orthree halogens.

The term “haloalkyl”, alone or in combination, denotes an alkyl groupsubstituted with at least one halogen, particularly substituted with oneto five halogens, particularly one to three halogens. A particular“haloalkyl” is trifluoromethyl.

The terms “hydroxyl” and “hydroxy”, alone or in combination, signify the—OH group.

The term “carbonyl”, alone or in combination, signifies the —C(O)—group.

The term “sulfonyl”, alone or in combination, signifies the —S(O)₂—group.

The term “pharmaceutically acceptable salts” refers to those salts whichretain the biological effectiveness and properties of the free bases orfree acids, which are not biologically or otherwise undesirable. Thesalts are formed with inorganic acids such as hydrochloric acid,hydrobromic acid, sulfuric acid, nitric acid, phosphoric acid,particularly hydrochloric acid, and organic acids such as acetic acid,propionic acid, glycolic acid, pyruvic acid, oxalic acid, maleic acid,malonic acid, succinic acid, fumaric acid, tartaric acid, citric acid,benzoic acid, cinnamic acid, mandelic acid, methanesulfonic acid,ethanesulfonic acid, p-toluenesulfonic acid, salicylic acid,N-acetylcystein. In addition these salts may be prepared form additionof an inorganic base or an organic base to the free acid. Salts derivedfrom an inorganic base include, but are not limited to, the sodium,potassium, lithium, ammonium, calcium, magnesium salts. Salts derivedfrom organic bases include, but are not limited to salts of primary,secondary, and tertiary amines, substituted amines including naturallyoccurring substituted amines, cyclic amines and basic ion exchangeresins, such as isopropylamine, trimethylamine, diethylamine,triethylamine, tripropylamine, ethanolamine, lysine, arginine,N-ethylpiperidine, piperidine, polyamine resins. The compound of formula(I) can also be present in the form of zwitterions. Particularlypreferred pharmaceutically acceptable salts of compounds of formula (I)are the salts of hydrochloric acid, hydrobromic acid, sulfuric acid,phosphoric acid and methanesulfonic acid.

“Pharmaceutically acceptable esters” means that the compound of generalformula (I) may be derivatised at functional groups to providederivatives which are capable of conversion back to the parent compoundsin vivo. Examples of such compounds include physiologically acceptableand metabolically labile ester derivatives, such as methoxymethylesters, methylthiomethyl esters and pivaloyloxymethyl esters.Additionally, any physiologically acceptable equivalents of the compoundof general formula (I), similar to the metabolically labile esters,which are capable of producing the parent compound of general formula(I) in vivo, are within the scope of this invention.

If one of the starting materials or compounds of formula (I) contain oneor more functional groups which are not stable or are reactive under thereaction conditions of one or more reaction steps, appropriateprotecting groups (as described e.g. in “Protective Groups in OrganicChemistry” by T. W. Greene and P. G. M. Wuts, 3^(rd) Ed., 1999, Wiley,New York) can be introduced before the critical step applying methodswell known in the art. Such protecting groups can be removed at a laterstage of the synthesis using standard methods described in theliterature. Examples of protecting groups are tert-butoxycarbonyl (Boc),9-fluorenylmethyl carbamate (Fmoc), 2-trimethylsilylethyl carbamate(Teoc), carbobenzyloxy (Cbz) and p-methoxybenzyloxycarbonyl (Moz).

The compound of formula (I) can contain several asymmetric centers andcan be present in the form of optically pure enantiomers, mixtures ofenantiomers such as, for example, racemates, mixtures ofdiastereoisomers, diastereoisomeric racemates or mixtures ofdiastereoisomeric racemates.

The term “asymmetric carbon atom” means a carbon atom with fourdifferent substituents. According to the Cahn-Ingold-Prelog Conventionan asymmetric carbon atom can be of the “R” or “S” configuration.

The invention relates in particular to:

A compound of formula (I) wherein A¹ is nitrogen;

A compound of formula (I) wherein A² is nitrogen;

A compound of formula (I) wherein A³ is —(CH₂)_(n)—;

A compound of formula (I) wherein A³ is —CH₂—;

A compound of formula (I) wherein R¹ is alkyl or cycloalkyl;

A compound of formula (I) wherein R¹ is alkyl;

A compound of formula (I) wherein R¹ is tert.-butyl or cyclopropyl;

A compound of formula (I) wherein R¹ is tert.-butyl;

A compound of formula (I) wherein R² is halopyrrolidinyl,hydroxypyrrolidinyl, alkoxy, halodihydropyrrolyl, alkylfurazanylalkoxy,hydroxyalkylpyrrolidinyl or alkoxyalkylpyrrolidinyl;

A compound of formula (I) wherein R² is difluoropyrrolidinyl,hydroxypyrrolidinyl, ethoxy, fluorodihydropyrrolyl,methylfurazanylmethoxy, hydroxymethylpyrrolidinyl ormethoxymethylpyrrolidinyl;

A compound of formula (I) wherein R² is substituted pyrrolidinyl,wherein substituted pyrrolidinyl is pyrrolidinyl substituted with one ortwo substituents independently selected from halogen and hydroxyl;

A compound of formula (I) wherein R² is difluoropyrrolidinyl orhydroxypyrrolidinyl;

A compound of formula (I) wherein R³ is phenyl, alkoxyphenyl,halophenyl, haloalkylphenyl, alkylfurazanyl, alkylsulfonylphenyl,pyridinyl, halopyridinyl, dioxothietanyl, tetrahydrofuranyl,alkyltetrazolyl, cycloalkyltetrazolyl, dialkyltriazolyl oralkyltriazolyl;

A compound of formula (I) wherein R³ is phenyl, methoxyphenyl,chlorophenyl, chlorofluorophenyl, trifluoromethylphenyl,methylfurazanyl, methylsulfonylphenyl, pyridinyl, chloropyridinyl,dioxothietanyl, tetrahydrofuranyl, methyltetrazolyl,cyclopropyltetrazolyl, dimethyltriazolyl or methyltriazolyl;

A compound of formula (I) wherein R³ is substituted phenyl, substitutedfurazanyl, substituted pyridinyl, substituted tetrazolyl or substitutedtriazolyl, wherein substituted phenyl, substituted pyridinyl andsubstituted triazolyl are phenyl, pyridinyl and triazolyl substitutedwith one or two substituents independently selected from alkyl, halogenand haloalkyl, wherein substituted tetrazolyl is tetrazolyl substitutedwith one substituent selected from alkyl, and cycloalkyl, and whereinsubstituted furazanyl is furazanyl substituted with alkyl;

A compound of formula (I) wherein R³ is trifluoromethylphenyl,methylfurazanyl, chloropyridinyl, methyltetrazolyl,cyclopropyltetrazolyl, dimethyltriazolyl or methyltriazolyl; and

A compound of formula (I) wherein n is 1.

The invention further relates in particular to a compound of formula (I)selected from

-   -   6-tert-Butyl-4-(3,3-difluoro-pyrrolidin-1-yl)-[(4-methoxyphenyl)methyl]-1H-pyrazolo[3,4-d]pyrimidine;    -   1-Benzyl-6-tert-butyl-4-(3,3-difluoro-pyrrolidin-1-yl)-1H-pyrazolo[3,4-d]pyrimidine;    -   (S)-1-[6-tert-Butyl-1-[(2-chlorophenyl)methyl]-1H-pyrazolo[3,4-d]pyrimidin-4-yl]-pyrrolidin-3-ol;    -   6-tert-Butyl-1-[(2-chlorophenyl)methyl]-4-(3,3-difluoro-pyrrolidin-1-yl)-1H-pyrazolo[3,4-d]pyrimidine;    -   6-tert-Butyl-1-[(2-chlorophenyl)methyl]-4-ethoxy-1H-pyrazolo[3,4-d]pyrimidine;    -   6-tert-Butyl-1-[(2-chloro-4-fluorophenyl)methyl]-4-(3,3-difluoro-pyrrolidin-1-yl)-1H-pyrazolo[3,4-d]pyrimidine;    -   6-tert-Butyl-4-(3,3-difluoro-pyrrolidin-1-yl)-1-[(2-trifluoromethylphenyl)methyl]-1H-pyrazolo[3,4-d]pyrimidine;    -   6-tert-Butyl-4-(3,3-difluoro-pyrrolidin-1-yl)-1-phenethyl-1H-pyrazolo[3,4-d]pyrimidine;    -   6-tert-Butyl-4-(3,3-difluoro-pyrrolidin-1-yl)-1-(4-methyl-furazan-3-ylmethyl)-1H-pyrazolo[3,4-d]pyrimidine;    -   6-tert-Butyl-4-(3,3-difluoro-pyrrolidin-1-yl)-1-(2-methanesulfonyl-benzyl)-1H-pyrazolo[3,4-d]pyrimidine;    -   6-tert-Butyl-4-(3,3-difluoro-pyrrolidin-1-yl)-1-(2-pyridin-3-yl-ethyl)-1H-pyrazolo[3,4-d]pyrimidine;    -   6-tert-Butyl-1-(2-chloro-pyridin-3-ylmethyl)-4-(3,3-difluoro-pyrrolidin-1-yl)-1H-pyrazolo[3,4-d]pyrimidine;    -   6-tert-Butyl-4-(3,3-difluoro-pyrrolidin-1-yl)-1-(1,1-dioxo-1λ6-thietan-3-yl)-1H-pyrazolo[3,4-d]pyrimidine;    -   6-tert-Butyl-4-(3,3-difluoro-pyrrolidin-1-yl)-1-(tetrahydro-furan-3-yl)-1H-pyrazolo[3,4-d]pyrimidine;    -   6-tert-Butyl-4-(3,3-difluoro-pyrrolidin-1-yl)-1-(1-methyl-1H-tetrazol-5-ylmethyl)-1H-pyrazolo[3,4-d]pyrimidine;    -   6-tert-Butyl-1-(1-cyclopropyl-1H-tetrazol-5-ylmethyl)-4-(3,3-difluoro-pyrrolidin-1-yl)-1H-pyrazolo[3,4-d]pyrimidine;    -   6-tert-Butyl-4-(3,3-difluoro-pyrrolidin-1-yl)-1-(4,5-dimethyl-4H-[1,2,4]triazol-3-ylmethyl)-1H-pyrazolo[3,4-d]pyrimidine;    -   6-tert-Butyl-1-(4,5-dimethyl-4H-[1,2,4]triazol-3-ylmethyl)-4-(3-fluoro-2,5-dihydro-pyrrol-1-yl)-1H-pyrazolo[3,4-d]pyrimidine;    -   6-tert-Butyl-4-(3,3-difluoro-pyrrolidin-1-yl)-1-(3-methyl-3H-[1,2,3]triazol-4-ylmethyl)-1H-pyrazolo[3,4-d]pyrimidine;    -   6-tert-Butyl-4-(3-fluoro-2,5-dihydro-pyrrol-1-yl)-1-(3-methyl-3H-[1,2,3]triazol-4-ylmethyl)-1H-pyrazolo[3,4-d]pyrimidine;    -   2-[6-tert-Butyl-4-(3,3-difluoro-pyrrolidin-1-yl)-pyrazolo[3,4-d]pyrimidin-1-yl]-1-pyridin-4-yl-ethanone;    -   2-[6-tert-Butyl-4-(3,3-difluoro-pyrrolidin-1-yl)-pyrazolo[3,4-d]pyrimidin-1-yl]-1-pyridin-2-yl-ethanone;    -   (S)-1-[6-tert-Butyl-1-[(4-methoxyphenyl)methyl]-1H-pyrazolo[3,4-d]pyrimidin-4-yl]-pyrrolidin-3-ol;    -   6-Cyclopropyl-4-(3,3-difluoro-pyrrolidin-1-yl)-1-[(4-methoxyphenyl)methyl]-1H-pyrazolo[3,4-b]pyridine;    -   (S)-1-[6-tert-Butyl-1-(4-methyl-furazan-3-ylmethyl)-1H-pyrazolo[3,4-d]pyrimidin-4-yl]-pyrrolidin-3-ol;    -   6-tert-Butyl-4-[(S)-3-(4-methyl-furazan-3-ylmethoxy)-pyrrolidin-1-yl]-1-(4-methyl-furazan-3-ylmethyl)-1H-pyrazolo[3,4-d]pyrimidine;    -   6-Cyclopropyl-4-(3,3-difluoro-pyrrolidin-1-yl)-1-(4-methyl-furazan-3-ylmethyl)-1H-pyrazolo[4,3-c]pyridine;    -   6-Cyclopropyl-4-(3,3-difluoro-pyrrolidin-1-yl)-1-(4-methyl-furazan-3-ylmethyl)-1H-pyrazolo[3,4-b]pyridine;    -   (S)-1-[6-tert-Butyl-1-[(2-trifluoromethylphenyl)methyl]-1H-pyrazolo[3,4-d]pyrimidin-4-yl]-pyrrolidin-3-ol;    -   (S)-1-[6-tert-Butyl-1-(2-methanesulfonyl-benzyl)-1H-pyrazolo[3,4-d]pyrimidin-4-yl]-pyrrolidin-3-ol;    -   (S)-1-[6-tert-Butyl-1-(1-methyl-1H-tetrazol-5-ylmethyl)-1H-pyrazolo[3,4-d]pyrimidin-4-yl]-pyrrolidin-3-ol;    -   (S)-1-[6-tert-Butyl-1-(3-chloro-pyridin-2-ylmethyl)-1H-pyrazolo[3,4-d]pyrimidin-4-yl]-pyrrolidin-3-ol;    -   (S)-1-[6-tert-Butyl-1-(1-cyclopropyl-1H-tetrazol-5-ylmethyl)-1H-pyrazolo[3,4-d]pyrimidin-4-yl]-pyrrolidin-3-ol;    -   {(R)-1-[6-tert-Butyl-1-[(2-trifluoromethylphenyl)methyl]-1H-pyrazolo[3,4-d]pyrimidin-4-yl]-pyrrolidin-2-yl}-methanol;    -   6-tert-Butyl-4-((R)-2-methoxymethyl-pyrrolidin-1-yl)-1-[(2-trifluoromethylphenyl)methyl]-1H-pyrazolo[3,4-d]pyrimidine;        and    -   6-tert-Butyl-4-((R)-2-methoxymethyl-pyrrolidin-1-yl)-1-(4-methyl-furazan-3-ylmethyl)-1H-pyrazolo[3,4-d]pyrimidine.

The invention further relates to a compound of formula (I) selected from

-   -   6-Chloro-4-(3,3-difluoropyrrolidin-1-yl)-1-[[2-(trifluoromethyl)phenyl]methyl]pyrazolo[3,4-d]pyrimidine;        and    -   4-(3,3-Difluoropyrrolidin-1-yl)-6-(2,2-dimethylpropoxy)-1-[[2-(trifluoromethyl)phenyl]methyl]pyrazolo[3,4-d]pyrimidine.

The invention also relates in particular to a compound of formula (I)selected from

-   -   6-tert-Butyl-4-(3,3-difluoro-pyrrolidin-1-yl)-1-[(2-trifluoromethylphenyl)methyl]-1H-pyrazolo[3,4-d]pyrimidine;    -   6-tert-Butyl-4-(3,3-difluoro-pyrrolidin-1-yl)-1-(4-methyl-furazan-3-ylmethyl)-1H-pyrazolo[3,4-d]pyrimidine;    -   6-tert-Butyl-1-(2-chloro-pyridin-3-ylmethyl)-4-(3,3-difluoro-pyrrolidin-1-yl)-1H-pyrazolo[3,4-d]pyrimidine;    -   6-tert-Butyl-4-(3,3-difluoro-pyrrolidin-1-yl)-1-(1-methyl-1H-tetrazol-5-ylmethyl)-1H-pyrazolo[3,4-d]pyrimidine;    -   6-tert-Butyl-1-(1-cyclopropyl-1H-tetrazol-5-ylmethyl)-4-(3,3-difluoro-pyrrolidin-1-yl)-1H-pyrazolo[3,4-d]pyrimidine;    -   6-tert-Butyl-4-(3,3-difluoro-pyrrolidin-1-yl)-1-(4,5-dimethyl-4H-[1,2,4]triazol-3-ylmethyl)-1H-pyrazolo[3,4-d]pyrimidine;    -   6-tert-Butyl-4-(3,3-difluoro-pyrrolidin-1-yl)-1-(3-methyl-3H-[1,2,3]triazol-4-ylmethyl)-1H-pyrazolo[3,4-d]pyrimidine;    -   (S)-1-[6-tert-Butyl-1-(4-methyl-furazan-3-ylmethyl)-1H-pyrazolo[3,4-d]pyrimidin-4-yl]-pyrrolidin-3-ol;        and    -   (S)-1-[6-tert-Butyl-1-[(2-trifluoromethylphenyl)methyl]-1H-pyrazolo[3,4-d]pyrimidin-4-yl]-pyrrolidin-3-ol.

The synthesis of the compound of formula (I) can, for example, beaccomplished according to the following schemes.

Unless otherwise specified, A¹ to A³, R¹ to R³ and n have in thefollowing schemes the meaning as defined above.

Following the procedure according to scheme 1, hydrazines of the generalformula AA can be used as starting material. Compounds AA are eithercommercially available, can be synthesized by a person skilled in theart as described in the literature, or obtained as described in theexperimental part.

Compound AB can be prepared from AA by reacting AA with2-(ethoxymethylen)-propanedinitrile (CAN 123-06-8) in the presence of abase, particularly DIEA, in an inert solvent, particularly ethanol attemperatures ranging from room temperature to the boiling point of themixture, preferably at 80-100° C.

Conversion of compound AB to AC can be achieved by hydrolysis in asuitable solvent, particularly by basic hydrolysis using a mixture ofpotassium hydroxide and hydrogen peroxide in dioxane-water mixtures atlow temperature, particularly at 0° C. to room temperature, or by usingother conditions known in the literature.

The acylation of the aromatic amine of general formula AC to givecompounds of formula AE can be affected by reaction with an acylatingagent AD. Acylating agents will preferably be acyl chlorides AD,particularly those that lack alpha hydrogen atoms next to the acyl groupto facilitate the following condensation step to compounds of type AF.The acylation itself is achieved by methods well known to the onesskilled in the art—using e.g. acyl chlorides AD in an inert solvent likeDMA, THF or mixtures thereof in the presence of a base like pyridine attemperatures between 0° C. and the reflux temperature of the solventemployed—preferably at 0° C. to room temperature.

Acyl chlorides AD are either commercially available, described in theliterature, can be synthesized by a person skilled in the art orobtained as described in the experimental part.

Dehydratisation and cyclisation of compounds AE gives compounds of thegeneral formula AF. This type of reaction is well known in the art andcan be done by heating compounds of type AE in the presence of a base,for example sodium hydroxide solution, either in the absence or in thepresence of an inert solvent, preferably by heating to elevatedtemperatures like 80° C. in the absence of additional solvents.

Compounds AF can be further elaborated to compounds AG by reaction withan inorganic acid chloride, for example phosphorus oxychloride in aninert, preferably high-boiling solvent, for example N,N-diethylanilineat elevated temperatures, for example 120° C.

Coupling a compound of formula AG with a nucleophile of formula AH togive compounds with the general structure I-a is conveniently done inthe presence of a base, like DIEA in an inert solvent, like DMF. Thereaction can for example be effected by heating the reaction mixture toelevated temperatures for example by heating to 120° C., preferably byreaction in a microwave at 120° C.

Nucleophiles AH are either commercially available, described in theliterature, can be synthesized by a person skilled in the art orobtained as described in the experimental part.

If one of the starting materials, compounds of formulae AA, AD or AH,contains one or more functional groups which are not stable or arereactive under the reaction conditions of one or more reaction steps,appropriate protecting groups (P) (as described e.g. in T. W. Greene etal., Protective Groups in Organic Chemistry, John Wiley and Sons Inc.New York 1999, 3^(rd) edition) can be introduced before the criticalstep applying methods well known in the art. Such protecting groups canbe removed at a later stage of the synthesis using standard methodsknown in the art.

If one or more compounds of formulae AA, AD or AH contain chiralcenters, compounds of formula I-a can be obtained as mixtures ofdiastereomers or enantiomers, which can be separated by methods wellknown in the art, e.g. (chiral) HPLC or crystallization. Racemiccompounds can e.g. be separated into their antipodes via diastereomericsalts by crystallization or by separation of the antipodes by specificchromatographic methods using either a chiral adsorbent or a chiraleluent.

Following the procedure according to scheme 2, compounds I-b (whereR^(3a)—CH₂ represents a suitable protecting group) can be used asstarting material for the synthesis of compounds I with other R¹ groups.

Compounds I-b (where R^(3a)—CH₂ represents a suitable protecting group)can be transformed to compounds BA by removal of the protecting group bymethods well known in the art, for example by hydrogenation or acidiccleavage reactions. A suitable sequence starts for example with a4-methoxybenzyl residue as R^(3a)—CH₂, that can be removed by acidicmethods for example by treatment with TFA and methanesulfonic acid in aninert solvent like DCM at temperatures from 0° C. to room temperature.

Compounds of the general formula I can be obtained from compounds of thegeneral formula BA by nucleophilic substitution reactions on compoundsof general formula BB (where X represents a leaving group on ansp3-carbon, as for example a halogen or pseudo-halogen). This reactioncan be effected by methods well known in the art, for example byreacting the partners in the presence of a base, like potassiumtert-butoxide, cesium carbonate or potassium carbonate in an inertsolvent like acetone, DMF or DMA, for example in a microwave oven atelevated temperatures like 120° C.

Compounds of formula BB are either commercially available, described inthe literature, can be synthesized by a person skilled in the art orobtained as described in the experimental part.

If one of the starting materials, compounds of formulae I-a or BB,contains one or more functional groups which are not stable or arereactive under the reaction conditions of one or more reaction steps,appropriate protecting groups (P) (as described e.g. in T. W. Greene etal., Protective Groups in Organic Chemistry, John Wiley and Sons Inc.New York 1999, 3^(rd) edition) can be introduced before the criticalstep applying methods well known in the art. Such protecting groups canbe removed at a later stage of the synthesis using standard methodsknown in the art.

If one or more compounds of formulae I-a or BB contain chiral centers,compounds of formula I-a can be obtained as mixtures of diastereomers orenantiomers, which can be separated by methods well known in the art,e.g. (chiral) HPLC or crystallization. Racemic compounds can e.g. beseparated into their antipodes via diastereomeric salts bycrystallization or by separation of the antipodes by specificchromatographic methods using either a chiral adsorbent or a chiraleluent.

Following the procedure according to scheme 3, substitutedaminopyrazoles of the general formula CA can be used as startingmaterial. Compounds CA are either commercially available, can besynthesized by a person skilled in the art as described in theliterature, or obtained as described in the experimental part.

Compound CB can be prepared from CA by reacting CA with diethyl malonate(CAN 105-53-3) either with or without an inert solvent, preferably inneat diethyl malonate at elevated temperature, preferably in a microwaveat 130° C.

Compounds CB can be further elaborated to compounds CC by reaction witha high boiling inorganic acid chloride, for example phenylphosphonicdichloride at elevated temperatures, for example 170° C.

Coupling a compound of formula CC with a nucleophile of formula AH togive compounds with the general structure CD is conveniently done in thepresence of a base, like DIEA in an inert solvent, like DMF. Thereaction can for example be effected by heating the reaction mixture toelevated temperatures for example by heating to 130° C., preferably byreaction in a microwave oven at 130° C.

Nucleophiles AH are either commercially available, described in theliterature, can be synthesized by a person skilled in the art orobtained as described in the experimental part.

Compounds I-c can be prepared from CD by coupling a compound of formulaCD with a suitably substituted alkyl, alkenyl or arylmetal species CE,particularly a cyclopropylboronic acid or cyclopropyltrifluoro-boratesalt in the presence of a suitable catalyst, particularly a palladiumcatalyst like palladium(II)acetate in the presence ofn-butyl-di-adamantylphosphine in an inert solvent such as toluene atroom temperature up to the reflux temperature of the solvent in thepresence of a suitable base, like cesium carbonate.

Alkyl, alkenyl or arylmetal species CE are either commerciallyavailable, described in the literature, can be synthesized by a personskilled in the art or obtained as described in the experimental part.

If one of the starting materials, compounds of formulae CA, AH or CE,contains one or more functional groups which are not stable or arereactive under the reaction conditions of one or more reaction steps,appropriate protecting groups (P) (as described e.g. in T. W. Greene etal., Protective Groups in Organic Chemistry, John Wiley and Sons Inc.New York 1999, 3^(rd) edition) can be introduced before the criticalstep applying methods well known in the art. Such protecting groups canbe removed at a later stage of the synthesis using standard methodsknown in the art.

If one or more compounds of formulae CA, AH or CE contain chiralcenters, compounds of formula I-c can be obtained as mixtures ofdiastereomers or enantiomers, which can be separated by methods wellknown in the art, e.g. (chiral) HPLC or crystallization. Racemiccompounds can e.g. be separated into their antipodes via diastereomericsalts by crystallization or by separation of the antipodes by specificchromatographic methods using either a chiral adsorbent or a chiraleluent.

Following the procedure according to scheme 4, compound DA(2,4,6-trichloro-3-pyridinecarboxaldehyde, CAN 1261269-66-2) can becondensed with hydrazines of general formula AA in the presence of abase, like DIEA and in an inert solvent, like THF at elevatedtemperatures, particularly at 50° C. to afford compounds of generalformula DB together with some regioisomers of DB.

Coupling a compound of formula DB with a nucleophile of formula AH togive compounds with the general structure DC is conveniently done in thepresence of a base, like DIEA in an inert solvent, like DMF. Thereaction can for example be affected by heating the reaction mixture toelevated temperatures for example by heating to 120° C., preferably byreaction in a microwave oven at 120° C.

Nucleophiles AH are either commercially available, described in theliterature, can be synthesized by a person skilled in the art orobtained as described in the experimental part.

Compounds I-d can be prepared from DC by coupling a compound of formulaDC with a suitably substituted alkyl, alkenyl or arylmetal species CE,particularly a cyclopropylboronic acid or cyclopropyltrifluoro-boratesalt in the presence of a suitable catalyst, particularly a palladiumcatalyst like palladium(II)acetate in the presence ofn-butyl-di-adamantylphosphine in an inert solvent such as toluene atroom temperature up to the reflux temperature of the solvent in thepresence of a suitable base, like cesium carbonate.

Alkyl, alkenyl or arylmetal species CE are either commerciallyavailable, described in the literature, can be synthesized by a personskilled in the art or obtained as described in the experimental part.

If one of the starting materials, compounds of formulae AA, AH or CE,contains one or more functional groups which are not stable or arereactive under the reaction conditions of one or more reaction steps,appropriate protecting groups (P) (as described e.g. in T. W. Greene etal., Protective Groups in Organic Chemistry, John Wiley and Sons Inc.New York 1999, 3^(rd) edition) can be introduced before the criticalstep applying methods well known in the art. Such protecting groups canbe removed at a later stage of the synthesis using standard methodsknown in the art.

If one or more compounds of formulae AA, AH or CE contain chiralcenters, compounds of formula I-d can be obtained as mixtures ofdiastereomers or enantiomers, which can be separated by methods wellknown in the art, e.g. (chiral) HPLC or crystallization. Racemiccompounds can e.g. be separated into their antipodes via diastereomericsalts by crystallization or by separation of the antipodes by specificchromatographic methods using either a chiral adsorbent or a chiraleluent.

The invention also relates to a process for the preparation of formula(I), comprising one of the following steps:

-   -   (a) the reaction of a compound of formula (A)

-   -   in the presence of R²H and a base;    -   (b) the reaction of a compound of formula (B)

-   -   in the presence of R³-A³-X and a base;    -   (c) the reaction of a compound of formula (C)

-   -   in the presence of MR¹, a palladium catalyst and a base;    -   (d) the reaction of a compound of formula (D)

-   -   in the presence of MR¹, a palladium catalyst and a base;    -   wherein A¹ to A³ and R¹ to R³ are as defined above, X is a        leaving group and M is a suitably substituted metalspecies, like        a borono-, a boryl-, a trifluoro-borate- or a stannyl species.

In step (a), the base is for example DIEA.

Step (a) is in particular advantageously carried out in an inertsolvent, like DMF. Step (a) can conveniently be carried out at 120° C.

In step (b), the base is for example tert.-butoxide, cesium carbonate orpotassium carbonate.

Step (b) is in particular advantageously carried out in an inertsolvent, like DMF or DMA. Step (b) can conveniently be carried out at120° C.

In step (c), the palladium catalyst is for example palladium(II)acetate.Step (c) is preferably carried out in the presence of a phosphine, inparticular n-butyl-di-adamantylphosphine.

Step (c) is in particular carried out in an inert solvent, such astoluene. Step (c) can conveniently be carried out at room temperature upto reflux.

In step (c), the base is for example cesium carbonate.

MR¹ is advantageously a cyclopropylboronic acid or acyclopropyltrifluoro-borate salt.

Similar conditions as for step (c) can advantageously be used for step(d).

If desired, the compound of formula (I) can be converted into apharmaceutically acceptable salt thereof.

The invention also relates in particular to:

The use of a compound of formula (I) for the treatment or prophylaxis ofpain, atherosclerosis, age-related macular degeneration, diabeticretinopathy, glaucoma, retinal vein occlusion, retinopathy ofprematurity, ocular ischemic syndrome, geographic atrophy, diabetesmellitus, inflammation, inflammatory bowel disease, ischemia-reperfusioninjury, acute liver failure, liver fibrosis, lung fibrosis, kidneyfibrosis, systemic fibrosis, acute allograft rejection, chronicallograft nephropathy, diabetic nephropathy, glomerulonephropathy,cardiomyopathy, heart failure, myocardial ischemia, myocardialinfarction, systemic sclerosis, thermal injury, burning, hypertrophicscars, keloids, gingivitis pyrexia, liver cirrhosis or tumors,regulation of bone mass, amyotrophic lateral sclerosis, multiplesclerosis, Alzheimer's disease, Parkinson's disease, stroke, transientischemic attack or uveitis;

The use of a compound according of formula (I) for the preparation of amedicament for the treatment or prophylaxis of pain, atherosclerosis,age-related macular degeneration, diabetic retinopathy, glaucoma,retinal vein occlusion, retinopathy of prematurity, ocular ischemicsyndrome, geographic atrophy, diabetes mellitus, inflammation,inflammatory bowel disease, ischemia-reperfusion injury, acute liverfailure, liver fibrosis, lung fibrosis, kidney fibrosis, systemicfibrosis, acute allograft rejection, chronic allograft nephropathy,diabetic nephropathy, glomerulonephropathy, cardiomyopathy, heartfailure, myocardial ischemia, myocardial infarction, systemic sclerosis,thermal injury, burning, hypertrophic scars, keloids, gingivitispyrexia, liver cirrhosis or tumors, regulation of bone mass, amyotrophiclateral sclerosis, multiple sclerosis, Alzheimer's disease, Parkinson'sdisease, stroke, transient ischemic attack or uveitis;

A compound of formula (I) for the treatment or prophylaxis of pain,atherosclerosis, age-related macular degeneration, diabetic retinopathy,glaucoma, retinal vein occlusion, retinopathy of prematurity, ocularischemic syndrome, geographic atrophy, diabetes mellitus, inflammation,inflammatory bowel disease, ischemia-reperfusion injury, acute liverfailure, liver fibrosis, lung fibrosis, kidney fibrosis, systemicfibrosis, acute allograft rejection, chronic allograft nephropathy,diabetic nephropathy, glomerulonephropathy, cardiomyopathy, heartfailure, myocardial ischemia, myocardial infarction, systemic sclerosis,thermal injury, burning, hypertrophic scars, keloids, gingivitispyrexia, liver cirrhosis or tumors, regulation of bone mass, amyotrophiclateral sclerosis, multiple sclerosis, Alzheimer's disease, Parkinson'sdisease, stroke, transient ischemic attack or uveitis; and

A method for the treatment or prophylaxis of pain, atherosclerosis,age-related macular degeneration, diabetic retinopathy, glaucoma,retinal vein occlusion, retinopathy of prematurity, ocular ischemicsyndrome, geographic atrophy, diabetes mellitus, inflammation,inflammatory bowel disease, ischemia-reperfusion injury, acute liverfailure, liver fibrosis, lung fibrosis, kidney fibrosis, systemicfibrosis, acute allograft rejection, chronic allograft nephropathy,diabetic nephropathy, glomerulonephropathy, cardiomyopathy, heartfailure, myocardial ischemia, myocardial infarction, systemic sclerosis,thermal injury, burning, hypertrophic scars, keloids, gingivitispyrexia, liver cirrhosis or tumors, regulation of bone mass, amyotrophiclateral sclerosis, multiple sclerosis, Alzheimer's disease, Parkinson'sdisease, stroke, transient ischemic attack or uveitis, which methodcomprises administering an effective amount of a compound of formula (I)to a patient in need thereof.

The invention particularly relates to a compound of formula (I) for thetreatment or prophylaxis of ischemia, reperfusion injury, liver fibrosisor kidney fibrosis, in particular ischemia or reperfusion injury.

The invention particularly relates to a compound of formula (I) for thetreatment or prophylaxis of myocardial infarction.

The invention further particularly relates to a compound of formula (I)for the treatment or prophylaxis of age-related macular degeneration,diabetic retinopathy, glaucoma, retinal vein occlusion, retinopathy ofprematurity, ocular ischemic syndrome, geographic atrophy or uveitis.

The invention further particularly relates to a compound of formula (I)for the treatment or prophylaxis of amyotrophic lateral sclerosis ormultiple sclerosis.

The invention is further directed to a compound of formula (I), whenmanufactured according to a process according to the invention.

Another embodiment of the invention provides a pharmaceuticalcomposition or medicament containing a compound of the invention and atherapeutically inert carrier, diluent or excipient, as well as a methodof using the compounds of the invention to prepare such composition andmedicament. In one example, the compound of formula (I) may beformulated by mixing at ambient temperature at the appropriate pH, andat the desired degree of purity, with physiologically acceptablecarriers, i.e., carriers that are non-toxic to recipients at the dosagesand concentrations employed into a galenical administration form. The pHof the formulation depends mainly on the particular use and theconcentration of compound, but preferably ranges anywhere from about 3to about 8. In one example, a compound of formula (I) is formulated inan acetate buffer, at pH 5. In another embodiment, the compound offormula (I) is sterile. The compound may be stored, for example, as asolid or amorphous composition, as a lyophilized formulation or as anaqueous solution.

Compositions are formulated, dosed, and administered in a fashionconsistent with good medical practice. Factors for consideration in thiscontext include the particular disorder being treated, the particularmammal being treated, the clinical condition of the individual patient,the cause of the disorder, the site of delivery of the agent, the methodof administration, the scheduling of administration, and other factorsknown to medical practitioners.

The compounds of the invention may be administered by any suitablemeans, including oral, topical (including buccal and sublingual),rectal, vaginal, transdermal, parenteral, subcutaneous, intraperitoneal,intrapulmonary, intradermal, intrathecal and epidural and intranasal,and, if desired for local treatment, intralesional administration.Parenteral infusions include intramuscular, intravenous, intraarterial,intraperitoneal, or subcutaneous administration.

The compounds of the present invention may be administered in anyconvenient administrative form, e.g., tablets, powders, capsules,solutions, dispersions, suspensions, syrups, sprays, suppositories,gels, emulsions, patches, etc. Such compositions may contain componentsconventional in pharmaceutical preparations, e.g., diluents, carriers,pH modifiers, sweeteners, bulking agents, and further active agents.

A typical formulation is prepared by mixing a compound of the presentinvention and a carrier or excipient. Suitable carriers and excipientsare well known to those skilled in the art and are described in detailin, e.g., Ansel, Howard C., et al., Ansel's Pharmaceutical Dosage Formsand Drug Delivery Systems. Philadelphia: Lippincott, Williams & Wilkins,2004; Gennaro, Alfonso R., et al. Remington: The Science and Practice ofPharmacy. Philadelphia: Lippincott, Williams & Wilkins, 2000; and Rowe,Raymond C. Handbook of Pharmaceutical Excipients. Chicago,Pharmaceutical Press, 2005. The formulations may also include one ormore buffers, stabilizing agents, surfactants, wetting agents,lubricating agents, emulsifiers, suspending agents, preservatives,antioxidants, opaquing agents, glidants, processing aids, colorants,sweeteners, perfuming agents, flavoring agents, diluents and other knownadditives to provide an elegant presentation of the drug (i.e., acompound of the present invention or pharmaceutical composition thereof)or aid in the manufacturing of the pharmaceutical product (i.e.,medicament).

The invention will now be illustrated by the following examples whichhave no limiting character.

EXAMPLES

Abbreviations

MS=mass spectrometry; EI=electron ionization; ESI=electrospray; NMR dataare reported in parts per million (δ) relative to internaltetramethylsilane and are referenced to the deuterium lock signal fromthe sample solvent (d₆-DMSO unless otherwise stated); coupling constants(J) are in Hertz, mp=melting point; bp=boiling point;DIEA=N-ethyl-N-isopropylpropan-2-amine;DBU=1,8-diazabicyclo[5.4.0]undec-7-ene; DCM=dichloromethane;DMA=dimethylacetamide; DMF=dimethylformamide; DMSO=dimethyl sulfoxide;dppf=1,1′-bis(diphenylphosphino)ferrocene;HATU=2-(3H-[1,2,3]triazolo[4,5-b]pyridin-3-yl)-1,1,3,3-tetramethylisouroniumhexafluorophosphate(V);HBTU=O-benzotriazole-N,N,N′,N′-tetramethyl-uronium-hexafluoro-phosphate;HPLC=LC=high performance liquid chromatography;m-CPBA=meta-chloroperoxybenzoic acid; Rt=retention time;TBAF=tetra-n-butylammonium fluoride;TBTU=O-(benzotriazol-1-yl)-N,N,N′,N′-tetramethyl-uronium-tetrafluoroborate;TEMPO=2,2,6,6-tetra-methylpiperidine 1-oxyl radical; TBME32 methyltert-butylether, THF=tetrahydrofuran; TFA=trifluoroacetic acid; tlc=thinlayer chromatography; CAN=CAS Registry Number.

Example 16-tert-Butyl-4-(3,3-difluoro-pyrrolidin-1-yl)-[(4-methoxyphenyl)methyl]-1H-pyrazolo[3,4-d]pyrimidinea) 5-Amino-1-[(4-methoxyphenyl)methyl]-1H-pyrazole-4-carbonitrile

[(4-Methoxyphenyl)methyl]hydrazine dihydrochloride (CAN 412327-07-2, 1.0g, 4.44 mmol), 2-(ethoxymethylene)-propanedinitrile (CAN 123-06-8, 0.542g, 4.44 mmol) and DIEA (1.55 mL, 8.88 mmol) were combined in ethanol (7mL) and the reaction mixture was stirred at 100° C. for 3 hours. Uponcooling in ice-water the product precipitated. Filtration, and dryinggave the title compound (0.597 g, 59%) as yellow solid; LC-MS (UV peakarea, ESI) 87%, 229.4 [MH⁺].

b) 5-Amino-1-[(4-methoxyphenyl)methyl]-1H-pyrazole-4-carboxamide

To a solution of potassium hydroxide (1.51 g, 27 mmol) in water (15 mL)were added at 0-5° C. hydrogen peroxide (3.64 mL, 119 mmol) and asolution of5-amino-1-[(4-methoxyphenyl)methyl]-1H-pyrazole-4-carbonitrile (0.55 g,2.4 mmol) in dioxane (30 mL). Cooling was removed and the reactionmixture was stirred at ambient temperature for 18 hours. The mixture waspartitioned between water (2×50 mL) and ethyl acetate; the organic phasewas separated, dried with Na₂SO₄, filtered and concentrated in vacuo.The crude material was purified by crystallization from ethylacetate/heptane to afford the desired product (0.51 g, 85%) aslight-yellow solid; LC-MS (UV peak area, ESI) 86%, 247.5 [MH⁺].

c)5-(2,2-Dimethyl-propionylamino)-1-[(4-methoxyphenyl)methyl]-1H-pyrazole-4-carboxylicacid amide

To a solution of5-amino-1-[(4-methoxyphenyl)methyl]-1H-pyrazole-4-carboxamide (400 mg,1.62 mmol) in DMA (4 mL) and THF (6 mL) was at 0° C. added pivaloylchloride (200 μL, 1.62 mmol) and pyridine (158 μL, 1.95 mmol) and themixture was stirred at 0° C. for 1.5 hours followed by stirring at roomtemperature for 2 hours. The mixture was partitioned between water (2×30mL) and DCM; the organic phase was separated, dried with Na₂SO₄,filtered and concentrated in vacuo. The crude material was purified byflash chromatography (silica, ethyl acetate/heptane gradient) to givethe desired product (1.07 g) as light-yellow oil that did still containsome DMA. The material was used in the next step without purification;LC-MS (UV peak area, ESI) 77%, 331.5 [MH⁺].

d)6-tert-Butyl-1,5-dihydro-1-[(4-methoxyphenyl)methyl]-4H-pyrazolo[3,4-d]pyrimidin-4-one

A solution of5-(2,2-dimethyl-propionylamino)-1-[(4-methoxyphenyl)methyl]-1H-pyrazole-4-carboxylicacid amide (0.54 g, 1.63 mmol) in sodium hydroxide solution (1 N, 10 mL)was stirred at 80° C. for 2.5 hours. After cooling, water (30 mL) wasadded and the mixture was extracted with DCM. Organic phases werepooled, dried with Na₂SO₄, filtered and concentrated in vacuo. The crudematerial was purified by flash chromatography (silica, ethylacetate/heptane gradient) to give the desired product (154 mg, 60% overtwo steps) as white solid; LC-MS (UV peak area, ESI) 95%, 313.5 [MH⁺].

e)6-tert-Butyl-4-chloro-1-[(4-methoxyphenyl)methyl]-1H-pyrazolo[3,4-d]pyrimidine

A mixture of6-tert-butyl-1,5-dihydro-1-[(4-methoxyphenyl)methyl]-4H-pyrazolo[3,4-d]pyrimidin-4-one(0.154 g, 0.49 mmol), phosphorus oxychloride (1.57 mL, 16.9 mmol) andN,N-diethylaniline (157 μL, 0.99 mmol) was stirred at 120° C. for 4.5hours. Phosphorus oxychloride was removed in vacuo and the residue waspartitioned between water and ethyl acetate; the organic phases werepooled, dried with Na₂SO₄, filtered and concentrated in vacuo to givethe desired product (144 mg) as dark green oil. The material was used inthe next step without purification.

f)6-tert-Butyl-4-(3,3-difluoro-pyrrolidin-1-yl)-1-(4-methoxy-benzyl)-1H-pyrazolo[3,4-d]pyrimidine

A mixture of6-tert-butyl-4-chloro-1-[(4-methoxyphenyl)methyl]-1H-pyrazolo[3,4-d]pyrimidine(0.144 g, 0.44 mmol), 3,3-difluoropyrrolidine hydrochloride (62.5 mg,0.44 mmol) and DIEA (380 μL, 2.18 mmol) in DMF (3 mL) was microwaved for1 hour at 120° C. After cooling, the mixture was partitioned betweenwater and ethyl acetate. Organic phases were pooled, dried with Na₂SO₄,filtered and concentrated in vacuo. The crude material was purified byflash chromatography (silica, ethyl acetate/heptane gradient) to givethe desired product (174 mg, 87% over two steps) as yellow oil; LC-MS(UV peak area, ESI) 90%, 402.6 [MH⁺].

Example 21-Benzyl-6-tert-butyl-4-(3,3-difluoro-pyrrolidin-1-yl)-1H-pyrazolo[3,4-d]pyrimidine

The title compound was synthesized in analogy to Examples 1c to 1f,using 5-amino-1-(phenylmethyl)-1H-pyrazole-4-carboxamide (CAN56156-22-0), pivaloyl chloride and 3,3-difluoropyrrolidine hydrochlorideas starting materials, and isolated (96 mg) as yellow solid; LC-MS (UVpeak area, ESI) 93%, 372.6 [MH⁺].

Example 3(S)-1-[6-tert-Butyl-1-[(2-chlorophenyl)methyl]-1H-pyrazolo[3,4-d]pyrimidin-4-yl]-pyrrolidin-3-ol

The title compound was synthesized in analogy to Examples 1c to 1f,using 5-amino-1-[(2-chlorophenyl)methyl]-1H-pyrazole-4-carboxamide (CAN106898-48-0), pivaloyl chloride and (S)-3-hydroxypyrrolidine (CAN100243-39-8) as starting materials, and isolated (2.5 mg) as colorlessoil; LC-MS (ESI) 386.5 [MH⁺].

Example 46-tert-Butyl-1-[(2-chlorophenyl)methyl]-4-(3,3-difluoro-pyrrolidin-1-yl)-1H-pyrazolo[3,4-d]pyrimidine

The title compound was synthesized in analogy to Examples 1c to 1f,using 5-amino-1-[(2-chlorophenyl)methyl]-1H-pyrazole-4-carboxamide (CAN106898-48-0), pivaloyl chloride and 3,3-difluoropyrrolidinehydrochloride as starting materials, and isolated (16 mg) as colorlessoil; LC-MS (UV peak area, ESI) 74%, 406.5 [MH⁺].

Example 56-tert-Butyl-1-[(2-chlorophenyl)methyl]-4-ethoxy-1H-pyrazolo[3,4-d]pyrimidine

The title compound was synthesized in analogy to Examples 1c to 1f,using 5-amino-1-[(2-chlorophenyl)methyl]-1H-pyrazole-4-carboxamide (CAN106898-48-0), pivaloyl chloride and ethanol as starting materials, andisolated (15 mg) as colorless oil; LC-MS (UV peak area, ESI) 83%, 345.5[MH⁺].

Example 66-tert-Butyl-1-[(2-chloro-4-fluorophenyl)methyl]-4-(3,3-difluoro-pyrrolidin-1-yl)-1H-pyrazolo[3,4-d]pyrimidinea)6-tert-Butyl-4-(3,3-difluoro-pyrrolidin-1-yl)-1H-pyrazolo[3,4-d]pyrimidine

To a mixture of6-tert-butyl-4-(3,3-difluoro-pyrrolidin-1-yl)-1-(4-methoxy-benzyl)-1H-pyrazolo[3,4-d]pyrimidine(0.174 g, 0.43 mmol) and TFA (870 μL, 11.3 mmol) in DCM (3 mL) was addedmethanesulfonic acid (141 μL, 2.17 mmol). The mixture was stirred for 1hour at 0° C. and for 1.5 hours at room temperature. Afterwards themixture was rendered basic with sodium hydroxide solution (2.5 mL, 25%,cooling) and partitioned between water and DCM. Organic phases werepooled, dried with MgSO₄, filtered and concentrated in vacuo. Theresidue, 135 mg of the title compound as a light-yellow solid, was usedwithout further purification; LC-MS (UV peak area, ESI) 80%, 282.5[MH⁺].

b)6-tert-Butyl-1-(2-chloro-4-fluoro-benzyl)-4-(3,3-difluoro-pyrrolidin-1-yl)-1H-pyrazolo[3,4-d]pyrimidine

To a solution of6-tert-butyl-4-(3,3-difluoro-pyrrolidin-1-yl)-1H-pyrazolo[3,4-d]pyrimidine(30 mg, 107 μmol) in acetone (1 mL) and DMF (2 mL) was added2-chloro-1-(chloromethyl)-4-fluoro-benzene (23 mg, 128 μmol) andpotassium carbonate (44 mg, 318 μmol). The mixture was microwaved for 45minutes at 120° C., cooled and partitioned between water and ethylacetate. Organic phases were pooled, dried by filtration over ChemElut®,and concentrated in vacuo. The residue was purified by preparative HPLC(Gemini NX, water/acetonitrile gradient), to afford the title compound(13 mg, 29%) as light-yellow oil; LC-MS (UV peak area, ESI) 78%, 424.5[MH⁺].

Example 76-tert-Butyl-4-(3,3-difluoro-pyrrolidin-1-yl)-1-[(2-trifluoromethylphenyl)methyl]-1H-pyrazolo[3,4-d]pyrimidine

The title compound was synthesized in analogy to Example 6 b, using6-tert-butyl-4-(3,3-difluoro-pyrrolidin-1-yl)-1H-pyrazolo[3,4-d]pyrimidine(Example 6 a) and 1-(chloromethyl)-2-(trifluoromethyl)-benzene (CAN21742-00-7) as starting materials, and isolated (7 mg, 14%) as colorlessoil; MS (ESI) 440.6 [MH⁺].

Example 86-tert-Butyl-4-(3,3-difluoro-pyrrolidin-1-yl)-1-phenethyl-1H-pyrazolo[3,4-d]pyrimidine

The title compound was synthesized in analogy to Example 6 b, using6-tert-butyl-4-(3,3-difluoro-pyrrolidin-1-yl)-1H-pyrazolo[3,4-d]pyrimidine(Example 6 a) and (2-bromoethyl)-benzene (CAN 103-63-9) as startingmaterials, and isolated (10 mg, 24%) as colorless oil; MS (ESI) 386.6[MH⁺].

Example 96-tert-Butyl-4-(3,3-difluoro-pyrrolidin-1-yl)-1-(4-methyl-furazan-3-ylmethyl)-1H-pyrazolo[3,4-d]pyrimidine

To a solution of6-tert-butyl-4-(3,3-difluoro-pyrrolidin-1-yl)-1H-pyrazolo[3,4-d]pyrimidine(Example 6 a, 30 mg, 107 μmol) in THF (0.3 mL) and DMA (0.5 mL) wasadded 3-(chloromethyl)-4-methyl-1,2,5-oxadiazole (28 mg, 213 μmol) andpotassium tert-butoxide (24 mg, 213 μmol). The mixture was microwavedfor 20 minutes at 110° C., cooled and partitioned between water andethyl acetate. Organic phases were pooled, dried by filtration overChemElut®, and concentrated in vacuo. The residue was purified bypreparative HPLC (Gemini NX, water/acetonitrile gradient), to afford thetitle compound (6 mg, 14%) as light-yellow oil; LC-MS (UV peak area,ESI) 94.9%, 378.1855 [MH⁺].

Example 106-tert-Butyl-4-(3,3-difluoro-pyrrolidin-1-yl)-1-(2-methanesulfonyl-benzyl)-1H-pyrazolo[3,4-d]pyrimidine

The title compound was synthesized in analogy to Example 9, using6-tert-butyl-4-(3,3-difluoro-pyrrolidin-1-yl)-1H-pyrazolo[3,4-d]pyrimidine(Example 6 a) and 1-(chloromethyl)-2-(methylsulfonyl)-benzene (CAN168551-51-7) as starting materials, and isolated (6 mg, 12%) ascolorless oil; LC-MS (UV peak area, ESI) 89.3%, 450.1776 [MH⁺].

Example 116-tert-Butyl-4-(3,3-difluoro-pyrrolidin-1-yl)-1-(2-pyridin-3-yl-ethyl)-1H-pyrazolo[3,4-d]pyrimidine

The title compound was synthesized in analogy to Example 9, using6-tert-butyl-4-(3,3-difluoro-pyrrolidin-1-yl)-1H-pyrazolo[3,4-d]pyrimidine(Example 6 a) and 2-(2-bromoethyl)-pyridine (CAN 39232-04-7) as startingmaterials, and isolated (6 mg, 12%) as colorless oil; MS (ESI) 387.6[MH⁺].

Example 126-tert-Butyl-1-(2-chloro-pyridin-3-ylmethyl)-4-(3,3-difluoro-pyrrolidin-1-yl)-1H-pyrazolo[3,4-d]pyrimidine

The title compound was synthesized in analogy to Example 9, using6-tert-butyl-4-(3,3-difluoro-pyrrolidin-1-yl)-1H-pyrazolo[3,4-d]pyrimidine(Example 6 a) and 2-chloro-3-(chloromethyl)-pyridine (CAN 89581-84-0) asstarting materials, and isolated (24 mg, 55%) as colorless oil; MS (ESI)407.5 [MH⁺].

Example 136-tert-Butyl-4-(3,3-difluoro-pyrrolidin-1-yl)-1-(1,1-dioxo-1λ⁶-thietan-3-yl)-1H-pyrazolo[3,4-d]pyrimidine

To a solution of6-tert-butyl-4-(3,3-difluoro-pyrrolidin-1-yl)-1H-pyrazolo[3,4-d]pyrimidine(Example 6 a, 30 mg, 107 μmol) in THF (0.3 mL) and DMF (0.3 mL) wasadded 3-chloro-thietane 1,1-dioxide (CAN 15953-83-0, 30 mg, 213 μmol),DIEA (33.5 μL, 192 μmol) and potassium tert-butoxide (24 mg, 213 μmol).The mixture was stirred for 1 hour at 0° C. and for 1 hour at roomtemperature and partitioned between water and ethyl acetate. Organicphases were pooled, dried by filtration over ChemElut®, and concentratedin vacuo. The residue was purified by preparative HPLC (Gemini NX,water/acetonitrile gradient), to afford the title compound (25 mg, 60%)as light-yellow oil; LC-MS (UV peak area, ESI) 97.7%, 386.1462 [MH⁺].

Example 146-tert-Butyl-4-(3,3-difluoro-pyrrolidin-1-yl)-1-(tetrahydro-furan-3-yl)-1H-pyrazolo[3,4-d]pyrimidine

The title compound was synthesized in analogy to Example 9, using6-tert-butyl-4-(3,3-difluoro-pyrrolidin-1-yl)-1H-pyrazolo[3,4-d]pyrimidine(Example 6 a) and 3-chlorotetrahydro-furan (CAN 19311-38-7) as startingmaterials, and isolated (1.4 mg, 4.5%) as yellow oil; MS (ESI) 352.4[MH⁺].

Example 156-tert-Butyl-4-(3,3-difluoro-pyrrolidin-1-yl)-1-(1-methyl-1H-tetrazol-5-ylmethyl)-1H-pyrazolo[3,4-d]pyrimidine

The title compound was synthesized in analogy to Example 9, using6-tert-butyl-4-(3,3-difluoro-pyrrolidin-1-yl)-1H-pyrazolo[3,4-d]pyrimidine(Example 6 a) and 5-(chloromethyl)-1-methyl-1H-tetrazole (CAN57235-84-4) as starting materials, and isolated (52 mg, 43%) as whitesolid; LC-MS (UV peak area, ESI) 100%, 378.1961 [MH⁺].

Example 166-tert-Butyl-1-(1-cyclopropyl-1H-tetrazol-5-ylmethyl)-4-(3,3-difluoro-pyrrolidin-1-yl)-1H-pyrazolo[3,4-d]pyrimidine

The title compound was synthesized in analogy to Example 9, using6-tert-butyl-4-(3,3-difluoro-pyrrolidin-1-yl)-1H-pyrazolo[3,4-d]pyrimidine(Example 6 a) and 5-(chloromethyl)-1-cyclopropyl-1H-tetrazole (CAN949980-56-7) as starting materials and cesium carbonate as base, andisolated (2 mg, 3.7%) as yellow oil; LC-MS (UV peak area, ESI) 91.8%,404.2114 [MH⁺].

Example 176-tert-Butyl-4-(3,3-difluoro-pyrrolidin-1-yl)-1-(4,5-dimethyl-4H-[1,2,4]triazol-3-ylmethyl)-1H-pyrazolo[3,4-d]pyrimidine

The title compound was synthesized in analogy to Example 9, using6-tert-butyl-4-(3,3-difluoro-pyrrolidin-1-yl)-1H-pyrazolo[3,4-d]pyrimidine(Example 6 a) and 3-(chloromethyl)-4,5-dimethyl-4H-1,2,4-triazole (CAN881845-16-5) as starting materials, and cesium carbonate as base, andisolated (2 mg, 4.8%) as yellow oil; LC-MS (UV peak area, ESI) 76.6%,391.2168 [MH⁺].

Example 186-tert-Butyl-1-(4,5-dimethyl-4H-[1,2,4]triazol-3-ylmethyl)-4-(3-fluoro-2,5-dihydro-pyrrol-1-yl)-1H-pyrazolo[3,4-d]pyrimidine

The title compound was isolated as side product during preparation ofExample 17, and isolated (4.9 mg, 12.4%) as yellow oil; LC-MS (UV peakarea, ESI) 91.7%, 371.2107 [MH⁺].

Example 196-tert-Butyl-4-(3,3-difluoro-pyrrolidin-1-yl)-1-(3-methyl-3H-[1,2,3]triazol-4-ylmethyl)-1H-pyrazolo[3,4-d]pyrimidine

The title compound was synthesized in analogy to Example 9, using6-tert-butyl-4-(3,3-difluoro-pyrrolidin-1-yl)-1H-pyrazolo[3,4-d]pyrimidine(Example 6 a) and 5-(chloromethyl)-1-methyl-1H-1,2,3-triazolehydrochloride (1:1) (CAN 327985-59-1) as starting materials, and cesiumcarbonate as base, and isolated (3 mg, 7.9%) as yellow oil; LC-MS (UVpeak area, ESI) 89.5%, 377.2007 [MH⁺].

Example 206-tert-Butyl-4-(3-fluoro-2,5-dihydro-pyrrol-1-yl)-1-(3-methyl-3H-[1,2,3]triazol-4-ylmethyl)-1H-pyrazolo[3,4-d]pyrimidine

The title compound was isolated as side product during preparation ofExample 17, and isolated (6 mg, 14.9%) as yellow oil; LC-MS (UV peakarea, ESI) 84.5%, 357.1948 [MH⁺].

Example 212-[6-tert-Butyl-4-(3,3-difluoro-pyrrolidin-1-yl)-pyrazolo[3,4-d]pyrimidin-1-yl]-1-pyridin-4-yl-ethanone

The title compound was synthesized in analogy to Example 9, using6-tert-butyl-4-(3,3-difluoro-pyrrolidin-1-yl)-1H-pyrazolo[3,4-d]pyrimidine(Example 6 a) and 2-bromo-1-(4-pyridinyl)-ethanone hydrobromide (1:1)(CAN 5349-17-7) as starting materials, and isolated (3.3 mg, 4.6%) asyellow oil; MS (ESI) 100%, 401.0 [MH⁺].

Example 222-[6-tert-Butyl-4-(3,3-difluoro-pyrrolidin-1-yl)-pyrazolo[3,4-d]pyrimidin-1-yl]-1-pyridin-2-yl-ethanone

The title compound was synthesized in analogy to Example 9, using6-tert-butyl-4-(3,3-difluoro-pyrrolidin-1-yl)-1H-pyrazolo[3,4-d]pyrimidine(Example 6 a) and 2-bromo-1-(2-pyridinyl)-ethanone hydrobromide (1:1)(CAN 17570-98-8) as starting materials, and isolated (2.0 mg, 2.8%) asbrown oil; MS (ESI) 100%, 401.0 [MH⁺].

Example 23(S)-1-[6-tert-Butyl-1-[(4-methoxyphenyl)methyl]-1H-pyrazolo[3,4-d]pyrimidin-4-yl]-pyrrolidin-3-ol

The title compound was synthesized in analogy to Example 1f, using6-tert-butyl-4-chloro-1-[(4-methoxyphenyl)methyl]-1H-pyrazolo[3,4-d]pyrimidine(Example 1e, 847 mg, 2.56 mmol) and (3S)-3-pyrrolidinol (CAN100243-39-8; 639 μL, 7.68 mmol) as starting materials and isolated (1.08g, quant.) as white foam; LC-MS (UV peak area, ESI) 96%, 382.7 [MH⁺].

Example 246-Cyclopropyl-4-(3,3-difluoro-pyrrolidin-1-yl)-1-[(4-methoxyphenyl)methyl]-1H-pyrazolo[3,4-b]pyridinea) 1-[(4-Methoxyphenyl)methyl]-1H-pyrazolo[3,4-b]pyridine-4,6-diol

1-[(4-Methoxyphenyl)methyl]-1H-pyrazol-5-amine (CAN 3528-45-8, 2.0 g,9.84 mmol), and diethyl malonate (25 mL, 164 mmol) were stirred for 15minutes at room temperature and subsequently warmed in a microwave ovenfor 3 hours to 130° C. After cooling the solid product was filtered offand dried to yield the title compound (1.25 g, 47%) as light yellowsolid; MS (ESI) 272.5 [MH⁺].

b) 4,6-Dichloro-1-[(4-methoxyphenyl)methyl]-1H-pyrazolo[3,4-b]pyridine

A mixture of1-[(4-methoxyphenyl)methyl]-1H-pyrazolo[3,4-b]pyridine-4,6-diol (0.9 g,3.32 mmol) and phenylphosphonic dichloride (7 mL, 49.9 mmol) was stirredat 170° C. for 20 hours. After cooling, the mixture was diluted with DCM(100 mL), basicified with 25% sodium hydroxide in ice water andpartitioned into DCM (3×100 mL); the organic phase was separated, driedwith MgSO₄, filtered and concentrated in vacuo. The crude material waspurified by flash chromatography (silica, ethyl acetate/DCM gradient) toafford the desired product (0.365 g, 36%) as light-yellow solid; MS(ESI) 308.4, 310.4 [MH⁺].

c)6-Chloro-4-(3,3-difluoro-pyrrolidin-1-yl)-1-[(4-methoxyphenyl)phenyl]-1H-pyrazolo[3,4-b]pyridine

To a solution of4,6-dichloro-1-[(4-methoxyphenyl)methyl]-1H-pyrazolo[3,4-b]pyridine (365mg, 1.18 mmol) in DMA (3 mL) was added 3,3-difluoropyrrolidinehydrochloride (340 mg, 2.37 mmol) and DIEA (2.07 mL, 11.8 mmol) and themixture was warmed in a microwave oven to 130° C. for 1 hour. Themixture was partitioned between water and TBME; the organic phases werepooled, dried with MgSO₄, filtered and concentrated in vacuo. The crudematerial was recrystallized from an ethyl acetate/heptane mixture togive the desired product (279 mg, 62%) as off-white solid; MS (ESI)379.5 [MH⁺].

d)6-Cyclopropyl-4-(3,3-difluoro-pyrrolidin-1-yl)-1-[(4-methoxyphenyl)methyl]-1H-pyrazolo[3,4-b]pyridine

To a solution of6-chloro-4-(3,3-difluoro-pyrrolidin-1-yl)-1-[(4-methoxyphenyl)phenyl]-1H-pyrazolo[3,4-b]pyridine(100 mg, 264 μmol), potassium cyclopropyltrifluoroborate (78.1 mg, 528μmol) and cesium carbonate (258 mg, 792 μmol) in toluene (1.55 mL) andwater (0.21 mL) was added Pd(OAc)₂ (5.9 mg, 26.4 μmol) andbutyl-bis(tricyclo[3.3.1.1^(3.7)]dec-1-yl)-phosphine (9.5 mg, 26.4μmol). The mixture was stirred at 100° C. for 6 hours and after cooling,passed through a Chemelut® cartridge and concentrated in vacuo. Thecrude material was purified by flash chromatography (silica, ethylacetate/heptane gradient) to give the desired product (36 mg, 31%) asyellow solid; MS (ESI) 385.6 [MH⁺].

Example 25(S)-1-[6-tert-Butyl-1-(4-methyl-furazan-3-ylmethyl)-1H-pyrazolo[3,4-d]pyrimidin-4-yl]-pyrrolidin-3-ola) (S)-1-(6-tert-Butyl-1H-pyrazolo[3,4-d]pyrimidin-4-yl)-pyrrolidin-3-ol

To a solution of(S)-1-[6-tert-butyl-1-(4-methoxy-benzyl)-1H-pyrazolo[3,4-d]pyrimidin-4-yl]-pyrrolidin-3-ol(Example 23, 890 mg, 2.33 mmol) in DCM (50 mL) was added at 0° C. TFA (3mL, 38.9 mmol) and methanesulfonic acid (0.6 mL, 9.24 mmol). The mixturewas kept in the fridge for 3 days and concentrated in vacuo. The residuewas partitioned between ethyl acetate and ice-cold sodium hydroxidesolution (25%, 10 mL). Organic phases were pooled, dried with Na₂SO₄,filtered and concentrated in vacuo. The crude material was purified byflash chromatography (silica, ethyl acetate/heptane gradient) to affordthe desired product (0.67 g, quant.) as white solid; LC-MS (UV peakarea, ESI) 98.2%, 262.1670 [MH⁺].

b)6-tert-Butyl-4-[(S)-3-(tert-butyl-dimethyl-silanyloxy)-pyrrolidin-1-yl]-1H-pyrazolo[3,4-d]pyrimidine

To a solution of(S)-1-(6-tert-butyl-1H-pyrazolo[3,4-d]pyrimidin-4-yl)-pyrrolidin-3-ol(576 mg, 2.2 mmol) and imidazole (300 mg, 4.41 mmol) in DMF (10 mL) wasadded a solution of tert-butyldimethylchlorosilane (664 mg, 4.41 mmol)in DMF (10 mL) during 30 minutes. The mixture was stirred at roomtemperature for 20 hours and afterwards concentrated in vacuo. Theresidue was partitioned between water (25 mL) and DCM (2×50 mL); theorganic phases were pooled, dried with MgSO₄, filtered and concentratedin vacuo. The crude material was purified by flash chromatography(silica, ethyl acetate/heptane gradient) to afford the desired product(0.445 g, 54%) as white solid; LC-MS (UV peak area, ESI) 99%, 376.6[MH⁺].

c)6-tert-Butyl-4-[(S)-3-(tert-butyl-dimethyl-silanyloxy)-pyrrolidin-1-yl]-1-(4-methyl-furazan-3-ylmethyl)-1H-pyrazolo[3,4-d]pyrimidine

To a solution of6-tert-butyl-4-[(S)-3-(tert-butyl-dimethyl-silanyloxy)-pyrrolidin-1-yl]-1H-pyrazolo[3,4-d]pyrimidine(70 mg, 186 μmol) in DMA (2 mL) was added3-(chloromethyl)-4-methyl-1,2,5-oxadiazole (39.5 mg, 238 μmol) andpotassium tert-butoxide (41.8 mg, 373 μmol). The mixture was microwavedfor 30 minutes at 150° C., cooled and partitioned between water andethyl acetate. Organic phases were pooled, dried by filtration overChemElut®, and concentrated in vacuo. The residue was purified bypreparative HPLC (Gemini NX, water/acetonitrile gradient), to afford thetitle compound (20 mg, 23%) as light-yellow oil; LC-MS (UV peak area,ESI) 99.6%, 472.2873 [MH⁺].

d)(S)-1-[6-tert-Butyl-1-(4-methyl-furazan-3-ylmethyl)-1H-pyrazolo[3,4-d]pyrimidin-4-yl]-pyrrolidin-3-ol

To a solution of6-tert-butyl-4-[(S)-3-(tert-butyl-dimethyl-silanyloxy)-pyrrolidin-1-yl]-1-(4-methyl-furazan-3-ylmethyl)-1H-pyrazolo[3,4-d]pyrimidine(20 mg, 42.4 μmol in THF (1 mL) was added TBAF (170 μL mg, 170 μmol).The mixture was stirred at room temperature for 2 hours and afterwards,passed through a Chemelut® cartridge and concentrated in vacuo. Thecrude material was purified by flash chromatography (silica, ethylacetate) to give the desired product (13 mg, 86%) as yellowish foam; MS(ESI) 358.7 [MH⁺].

Example 266-tert-Butyl-4-[(S)-3-(4-methyl-furazan-3-ylmethoxy)-pyrrolidin-1-yl]-1-(4-methyl-furazan-3-ylmethyl)-1H-pyrazolo[3,4-d]pyrimidine

The title compound was isolated as side-product in Example 26 c (4.4 mg,5.2%) as colorless wax; LC-MS (UV peak area, ESI) 93.2%, 454.2312 [MH⁺].

Example 276-Cyclopropyl-4-(3,3-difluoro-pyrrolidin-1-yl)-1-(4-methyl-furazan-3-ylmethyl)-1H-pyrazolo[4,3-c]pyridinea) 4,6-Dichloro-1-[(4-methoxyphenyl)methyl]-1H-pyrazolo[4,3-c]pyridine

To a solution of 2,4,6-trichloro-3-pyridinecarboxaldehyde (CAN1261269-66-2, 375 mg, 178 mmol) and DIEA (1.56 mL, 8.91 mmol) in THF(4.0 mL) at 50° C. was added with stirring a solution of[(4-methoxyphenyl)methyl]-hydrazine hydrochloride (1:1) (370 mg, 1.96mmol) in THF (4.0 mL). The mixture was stirred for 20 minutes at 50° C.,cooled and partitioned between ethyl acetate and water. Organic phaseswere pooled, dried with Na₂SO₄, filtered and concentrated in vacuo. Thecrude material was purified by flash chromatography (silica, ethylacetate/heptane gradient) to afford the desired product (0.21 g, 38%) aswhite solid; LC-MS (UV peak area, ESI) 86.4%, 308.0358, 310.0328 [MH⁺].

b)6-Chloro-4-(3,3-difluoro-pyrrolidin-1-yl)-1-[(4-methoxyphenyl)methyl]-1H-pyrazolo[4,3-c]pyridine

To a solution of4,6-dichloro-1-[(4-methoxyphenyl)methyl]-1H-pyrazolo[4,3-c]pyridine (185mg, 0.6 mmol) and DIEA (839 μL, 4.8 mmol) in DMF (2 mL) was added3,3-difluoropyrrolidine hydrochloride (129 mg, 0.9 mmol). The mixturewas warmed in the microwave oven for 1 hour at 120° C., cooled andpartitioned between water and ethyl acetate. The organic phases werepooled, dried with MgSO₄, filtered and concentrated in vacuo. The crudematerial was purified by flash chromatography (silica, ethylacetate/heptane gradient) to afford the desired product (186 mg, 82%) asyellow solid; LC-MS (UV peak area, ESI) 97.4%, 379.1133 [MH⁺].

c)6-Cyclopropyl-4-(3,3-difluoro-pyrrolidin-1-yl)-1-[(4-methoxyphenyl)methyl]-1H-pyrazolo[4,3-c]pyridine

To a solution of6-chloro-4-(3,3-difluoro-pyrrolidin-1-yl)-1-[(4-methoxyphenyl)methyl]-1H-pyrazolo[4,3-c]pyridine(120 mg, 317 μmol), potassium cyclopropyltrifluoroborate (141 mg, 950μmol) and cesium carbonate 310 mg, 950 μmol) in toluene (0.8 mL) andwater (0.1 mL) was added Pd(OAc)₂ (7.1 mg, 31.7 μmol) andbutyl-bis(tricyclo[3.3.1.1^(3,7)]dec-1-yl)-phosphine (11.4 mg, 31.7μmol). The mixture was stirred at 110° C. for 5 hours and after cooling,passed through a Chemelut® cartridge and concentrated in vacuo. Thecrude material was purified by flash chromatography (silica, ethylacetate/heptane gradient) to give the desired product (116 mg, 85%) asyellow wax; MS (ESI) 385.6 [MH⁺].

d)6-Cyclopropyl-4-(3,3-difluoro-pyrrolidin-1-yl)-1H-pyrazolo[4,3-c]pyridine

To a solution of6-cyclopropyl-4-(3,3-difluoro-pyrrolidin-1-yl)-1-[(4-methoxyphenyl)methyl]-1H-pyrazolo[4,3-c]pyridine(116 mg, 302 μmol) in DCM (1.5 mL) was added at 0° C. TFA (395 μL, 5.1mmol) and methanesulfonic acid (78.4 μL, 1.21 mmol). The mixture wasstirred at 0° C. for 1 hour, at room temperature overnight and for 3hours at 40° C. After cooling 25% sodium hydroxide in ice water wasadded, the mixture was dried by extraction over Chemelut® andconcentrated in vacuo. The crude material was purified by flashchromatography (silica, ethyl acetate/heptane gradient) to afford thedesired product (81 mg, quant.) as brown solid; MS (ESI) 265.6 [MH⁺].

e)6-Cyclopropyl-4-(3,3-difluoro-pyrrolidin-1-yl)-1-(4-methyl-furazan-3-ylmethyl)-1H-pyrazolo[4,3-c]pyridine

The title compound was synthesized in analogy to Example 9, using6-cyclopropyl-4-(3,3-difluoro-pyrrolidin-1-yl)-1H-pyrazolo[4,3-c]pyridine(Example 27 d, 81 mg, 307 μmol) and3-(chloromethyl)-4-methyl-1,2,5-oxadiazole (81.3 mg, 613 μmol) asstarting materials, and isolated (23 mg, 21%) as yellow oil; LC-MS (UVpeak area, ESI) 99.2%, 361.1591 [MH⁺].

Example 286-Cyclopropyl-4-(3,3-difluoro-pyrrolidin-1-yl)-1-(4-methyl-furazan-3-ylmethyl)-1H-pyrazolo[3,4-b]pyridinea)6-Cyclopropyl-4-(3,3-difluoro-pyrrolidin-1-yl)-1H-pyrazolo[3,4-b]pyridine

The title compound was synthesized in analogy to Example 27 d, using6-cyclopropyl-4-(3,3-difluoro-pyrrolidin-1-yl)-1-[(4-methoxyphenyl)methyl]-1H-pyrazolo[3,4-b]pyridine(Example 24 d, 90 mg, 234 μmol) as starting material, and isolated (63mg, quant.) as yellow oil; and used without further characterization inthe next step.

b)6-Cyclopropyl-4-(3,3-difluoro-pyrrolidin-1-yl)-1-(4-methyl-furazan-3-ylmethyl)-1H-pyrazolo[3,4-b]pyridine

The title compound was synthesized in analogy to Example 9, using6-cyclopropyl-4-(3,3-difluoro-pyrrolidin-1-yl)-1H-pyrazolo[3,4-b]pyridine(Example 28 a, 79 mg, 299 μmol) and3-(chloromethyl)-4-methyl-1,2,5-oxadiazole (79.2 mg, 598 μmol) asstarting materials, and isolated (5.6 mg, 4.2%) as brown oil; LC-MS (UVpeak area, ESI) 95.1%, 361.1585 [MH⁺].

Example 29(S)-1-[6-tert-Butyl-1-[(2-trifluoromethylphenyl)methyl]-1H-pyrazolo[3,4-d]pyrimidin-4-yl]pyrrolidin-3-ola)5-Amino-1-[(2-trifluoromethylphenyl)methyl]-1H-pyrazole-4-carbonitrile

The title compound was synthesized in analogy to Example 1 a, using[[2-(trifluoromethyl)phenyl]methyl]-hydrazine hydrochloride (1:1) (CAN1263378-37-5, 1.0 g, 4.28 mmol) and 2-(ethoxymethylene)-propanedinitrile(CAN 123-06-8, 533 mg, 4.28 mmol) as starting materials, and isolated(855 mg, 75%) as yellow solid; MS (ESI) 267.5 [MH⁺].

b) 5-Amino-1-[(2-trifluoromethylphenyl)methyl]-1H-pyrazole-4-carboxamide

The title compound was synthesized in analogy to Example 1 b, using5-amino-1-[(2-trifluoromethylphenyl)methyl]-1H-pyrazole-4-carbonitrile(Example 29 a, 850 mg, 3.19 mmol) as starting material, and isolated(725 mg, 80%) as light yellow solid; MS (ESI) 285.5 [MH⁺].

c)5-(2,2-Dimethyl-propionylamino)-1-[(2-trifluoromethylphenyl)methyl]-1H-pyrazole-4-carboxylicacid amide

The title compound was synthesized in analogy to Example 1 c, using5-amino-1-[(2-trifluoromethylphenyl)methyl]-1H-pyrazole-4-carboxamide(Example 29 b, 720 mg, 2.53 mmol) and pivaloyl chloride (312 μL, 2.53mmol) as starting materials, and isolated (0.95 g, quant.) as lightyellow oil; MS (ESI) 396.6 [MH⁺].

d)6-tert-Butyl-1,5-dihydro-1-[(2-trifluoromethylphenyl)methyl]-4H-pyrazolo[3,4-d]pyrimidin-4-one

The title compound was synthesized in analogy to Example 1 d, using5-(2,2-dimethyl-propionylamino)-1-[(2-trifluoromethylphenyl)methyl]-1H-pyrazole-4-carboxylicacid amide (Example 29 c, 933 mg, 3.19 mmol) as starting material, andisolated (408 mg, 80%) as white solid; MS (ESI) 351.6 [MH⁺].

e)6-tert-Butyl-4-chloro-1-[(2-trifluoromethylphenyl)methyl]-1H-pyrazolo[3,4-d]pyrimidine

The title compound was synthesized in analogy to Example 1 e, using6-tert-butyl-1,5-dihydro-1-[(2-trifluoromethylphenyl)methyl]-4H-pyrazolo[3,4-d]pyrimidin-4-one(Example 29 d, 200 mg, 571 μmol) as starting material, and isolated (458mg, quant.) as black oil that was used in the next step without furthercharacterization.

f)(S)-1-[6-tert-Butyl-1-[(2-trifluoromethylphenyl)methyl]-1H-pyrazolo[3,4-d]pyrimidin-4-yl]-pyrrolidin-3-ol

The title compound was synthesized in analogy to Example 1 f, using6-tert-butyl-4-chloro-1-[(2-trifluoromethylphenyl)methyl]-1H-pyrazolo[3,4-d]pyrimidine(Example 29 d, 211 mg, 572 μmol) and (3S)-3-pyrrolidinol (CAN100243-39-8, 143 μL, 1.72 mmol) as starting materials, and isolated (124mg, 42%) as white foam; LC-MS (UV peak area; ESI) 100%, 420.2022 [MH⁺].

Example 30(S)-1-[6-tert-Butyl-1-(2-methanesulfonyl-benzyl)-1H-pyrazolo[3,4-d]pyrimidin-4-yl]-pyrrolidin-3-ol

The title compound was synthesized in analogy to Example 9, using6-tert-butyl-4-[(S)-3-(tert-butyl-dimethyl-silanyloxy)-pyrrolidin-1-yl]-1H-pyrazolo[3,4-d]pyrimidine(Example 25 b, 30 mg, 80 μmol) and1-(chloromethyl)-2-(methylsulfonyl)-benzene (CAN 168551-51-7; 33 mg, 160μmol) as starting materials and isolated (2 mg, 5.8%) as colorless oil;LC-MS (UV peak area, ESI) 87.4%, 430.1911 [MH⁺].

Example 31(S)-1-[6-tert-Butyl-1-(1-methyl-1H-tetrazol-5-ylmethyl)-1H-pyrazolo[3,4-d]pyrimidin-4-yl]-pyrrolidin-3-ol

The title compound was synthesized in analogy to Example 9, using6-tert-butyl-4-[(S)-3-(tert-butyl-dimethyl-silanyloxy)-pyrrolidin-1-yl]-1H-pyrazolo[3,4-d]pyrimidine(Example 25 b, 80 mg, 213 μmol) and5-(chloromethyl)-1-methyl-1H-tetrazole (CAN 57235-84-4; 38 mg, 287 μmol)as starting materials and isolated (12 mg, 16%) as colorless oil; LC-MS(UV peak area, ESI) 93%, 358.7 [MH⁺].

Example 32(S)-1-[6-tert-Butyl-1-(3-chloro-pyridin-2-ylmethyl)-1H-pyrazolo[3,4-d]pyrimidin-4-yl]-pyrrolidin-3-ol

The title compound was synthesized in analogy to Example 9, using6-tert-butyl-4-[(S)-3-(tert-butyl-dimethyl-silanyloxy)-pyrrolidin-1-yl]-1H-pyrazolo[3,4-d]pyrimidine(Example 25 b, 60 mg, 160 μmol) and 3-chloro-2-(chloromethyl)-pyridine(CAN 185315-53-1; 51.8 mg, 320 μmol) as starting materials and after anadditional deprotection step with TBAF isolated (13 mg, 21%) ascolorless oil; LC-MS (UV peak area, ESI) 100%, 387.1703 [MH⁺].

Example 33(S)-1-[6-tert-Butyl-1-(1-cyclopropyl-1H-tetrazol-5-ylmethyl)-1H-pyrazolo[3,4-d]pyrimidin-4-yl]-pyrrolidin-3-ol

The title compound was synthesized in analogy to Example 9, using6-tert-butyl-4-[(S)-3-(tert-butyl-dimethyl-silanyloxy)-pyrrolidin-1-yl]-1H-pyrazolo[3,4-d]pyrimidine(Example 25 b, 120 mg, 320 μmol) and5-(chloromethyl)-1-cyclopropyl-1H-tetrazole (CAN 949980-56-7; 76 mg, 479μmol) as starting materials and after an additional deprotection stepwith TBAF isolated (30 mg, 26%) as white foam; LC-MS (UV peak area, ESI)97%, 384.7 [MH⁺].

Example 34

{(R)-1-[6-tert-Butyl-1-[(2-trifluoromethylphenyl)methyl]-1H-pyrazolo[3,4-d]pyrimidin-4-yl]pyrrolidin-2-yl}-methanol

The title compound was synthesized in analogy to Example 1 f, using6-tert-butyl-4-chloro-1-[(2-trifluoromethylphenyl)methyl]-1H-pyrazolo[3,4-d]pyrimidine(Example 29 d, 100 mg, 271 μmol) and (2R)-2-pyrrolidinemethanol (CAN68832-13-3, 82.3 mg, 813 μmol) as starting materials, and isolated (110mg, 94%) as colorless oil; LC-MS (UV peak area; ESI) 98%, 434.7 [MH⁺].

Example 356-tert-Butyl-4-((R)-2-methoxymethyl-pyrrolidin-1-yl)-1-[(2-trifluoromethylphenyl)methyl]-1H-pyrazolo[3,4-d]pyrimidine

The title compound was synthesized in analogy to Example 1 f, using6-tert-butyl-4-chloro-1-[(2-trifluoromethylphenyl)methyl]-1H-pyrazolo[3,4-d]pyrimidine(Example 29 d, 100 mg, 271 μmol) and (2R)-2-(methoxymethyl)-pyrrolidine(CAN 84025-81-0, 93.7 mg, 813 μmol) as starting materials, and isolated(106 mg, 87%) as colorless oil; LC-MS (UV peak area; ESI) 95%, 448.7[MH⁺].

Example 366-tert-Butyl-4-((R)-2-methoxymethyl-pyrrolidin-1-yl)-1-(4-methyl-furazan-3-ylmethyl)-1H-pyrazolo[3,4-d]pyrimidinea)(1,3-Dioxo-1,3-dihydro-isoindol-2-yl)-(4-methyl-furazan-3-ylmethyl)-carbamicacid tert-butyl ester

To a mixture of N-(1,3-dihydro-1,3-dioxo-2H-isoindol-2-yl)-carbamic acid1,1-dimethylethyl ester (CAN 34387-89-8, 5.0 g, 19.1 mmol), potassiumcarbonate (10.5 g, 76.3 mmol) and benzyl triethylammonium chloride (0.65g, 2.86 mmol) in acetonitrile (150 mL) was added3-(bromomethyl)-4-methyl-1,2,5-oxadiazole (CAN 90507-32-7, 4.05 g, 22.9mmol). The mixture was stirred at room temperature for 10 minutes, overnight at 60° C. and subsequently filtered. The filtrate was concentratedin vacuo and the crude material was purified by flash chromatography(silica, ethyl acetate/heptane gradient) to afford the desired product(5.52 g, 81%) as white solid; LC-MS (UV peak area; ESI) 99%, 257.4[M-C₄H₉CO₂ ⁻].

b) N-(4-Methyl-furazan-3-ylmethyl)-hydrazinecarboxylic acid tert-butylester

To a solution of(1,3-dioxo-1,3-dihydro-isoindol-2-yl)-(4-methyl-furazan-3-ylmethyl)-carbamicacid tert-butyl ester (4.5 g, 12.6 mmol) in ethanol (50 mL) and THF (30mL) was added hydrazine (2.0 mL, 63.7 mmol). The mixture was stirredover night at 70° C. and subsequently filtered. The filtrate wasconcentrated in vacuo and the crude title compound (3.0 g, quant.) wasused in subsequent steps without further purification; GC-MS (TIC peakarea; EI) 95%, 228 [M].

c) (4-Methyl-furazan-3-ylmethyl)-hydrazine hydrochloride (1:1)

To a solution of N-(4-methyl-furazan-3-ylmethyl)-hydrazinecarboxylicacid tert-butyl ester (3.0 g, 13.1 mmol) in methanol (200 mL) was addedhydrochloric acid in dioxane (4 N, 4 mL). The mixture was stirred atroom temperature for 5 days at room temperature and concentrated invacuo. The crude material was purified by recrystallization from ethylacetate to afford the desired product in ˜85% purity (2.1 g, 84%) aswhite solid; LC-MS (ESI) 129.0767 [MH⁺].

d) 5-Amino-1-(4-methyl-furazan-3-ylmethyl)-1H-pyrazole-4-carbonitrile

The title compound was synthesized in analogy to Example 1 a, using(4-methyl-furazan-3-ylmethyl)-hydrazine hydrochloride (1:1) (Example 26c, 0.49 g, 2.97 mmol) and 2-(ethoxymethylene)-propanedinitrile (CAN123-06-8, 533 mg, 3.27 mmol) as starting materials, and isolated (298mg, 49%) as yellow solid; MS (ESI) 205.5 [MH⁺].

e) 5-Amino-1-(4-methyl-furazan-3-ylmethyl)-1H-pyrazole-4-carboxylic acidamide

The title compound was synthesized in analogy to Example 1 b, using5-amino-1-(4-methyl-furazan-3-ylmethyl)-1H-pyrazole-4-carbonitrile(Example 36 2, 298 mg, 1.47 mmol) as starting material, and isolated(310 mg, 85%) as yellow solid; MS (ESI) 223.2 [MH⁺].

f)5-(2,2-Dimethyl-propionylamino)-1-(4-methyl-furazan-3-ylmethyl)-1H-pyrazole-4-carboxylicacid amide

The title compound was synthesized in analogy to Example 1 c, using5-amino-1-(4-methyl-furazan-3-ylmethyl)-1H-pyrazole-4-carboxylic acidamide (Example 36 e, 360 mg, 1.62 mmol) and pivaloyl chloride (199 μL,1.62 mmol) as starting materials, and isolated (0.49 g, quant.) asyellow oil; MS (ESI) 307.4 [MH⁺].

g)6-tert-Butyl-1-(4-methyl-furazan-3-ylmethyl)-1,5-dihydro-pyrazolo[3,4-d]pyrimidin-4-one

The title compound was synthesized in analogy to Example 1 d, using5-(2,2-dimethyl-propionylamino)-1-(4-methyl-furazan-3-ylmethyl)-1H-pyrazole-4-carboxylicacid amide (Example 36 f, 107 mg, 0.35 mmol) as starting material, andisolated (88 mg, 87%) as white solid; MS (ESI) 289.5 [MH⁺].

h)6-tert-Butyl-4-chloro-1-(4-methyl-furazan-3-ylmethyl)-1H-pyrazolo[3,4-d]pyrimidine

The title compound was synthesized in analogy to Example 1 e, using6-tert-butyl-1-(4-methyl-furazan-3-ylmethyl)-1,5-dihydro-pyrazolo[3,4-c]pyrimidin-4-one(Example 36 g, 135 mg, 468 μmol) as starting material, and isolated (144mg, quant.) as yellow oil that was used in the next step without furthercharacterization.

i)6-tert-Butyl-4-((R)-2-methoxymethyl-pyrrolidin-1-yl)-1-(4-methyl-furazan-3-ylmethyl)-1H-pyrazolo[3,4-c]pyrimidine

The title compound was synthesized in analogy to Example 1 f, using6-tert-butyl-4-chloro-1-(4-methyl-furazan-3-ylmethyl)-1H-pyrazolo[3,4-c]pyrimidine(Example 36 h, 144 mg, 468 μmol) and (2R)-2-(methoxymethyl)-pyrrolidine(CAN 84025-81-0, 93.3 mg, 810 μmol) as starting materials, and isolated(125 mg, 69%) as brown oil; LC-MS (UV peak area; ESI) 98.5%, 386.2311[MH⁺].

Example 376-Chloro-4-(3,3-difluoropyrrolidin-1-yl)-1-[[2-(trifluoromethyl)phenyl]methyl]pyrazolo[3,4-d]pyrimidinea)4,6-Dichloro-1-(2-(trifluoromethyl)benzyl)-1H-pyrazolo[3,4-d]pyrimidine

A suspension of5-amino-1-[(2-trifluoromethylphenyl)methyl]-1H-pyrazole-4-carboxamide(2500 mg, 8.8 mmol, Example 29 b), diethyl carbonate (1.25 g, 1.28 mL,10.6 mmol) and sodium tert-butoxide (1.69 g, 17.6 mmol) in ethanol (70mL) and DMSO (10 mL) was stirred for 8 d at 100° C. The yellowsuspension was cooled to 0° C. and filtrated. The residue washed withtBuOMe and the combined filtrates were concentrated in vacuo. Heptanewas added to precipitate the title compound (2.4 g, 84%) as white solidwhich was used in the next step without further purification; MS (ESI)311.5 [MH⁺].

b)4,6-Dichloro-1-(2-(trifluoromethyl)benzyl)-1H-pyrazolo[3,4-d]pyrimidine

A mixture of1-(2-(trifluoromethyl)benzyl)-1H-pyrazolo[3,4-d]pyrimidine-4,6-diol (500mg, 1.61 mmol) and POCl₃ (12.4 g, 7.51 mL, 80.6 mmol) andN,N-diethylaniline (433 mg, 464 μL, 2.9 mmol) was heated under stirringfor 2 h to 110° C. The crude reaction mixture was concentrated in vacuo,poured into 25 mL dichloromethane and washed with ice water (2×25 mL).The combined aqueous layers were back-extracted with dichloromethane(1×25 mL). The combined organic layers were dried over MgSO₄ andconcentrated in vacuo. The crude material was purified by chromatography(silica gel, 20 g, AcOEt/heptane 1/3) to give the title compound (50 mg,9%) as white solid; MS (ESI) 347.4 [MH⁺].

c)6-Chloro-4-(3,3-difluoropyrrolidin-1-yl)-1-[[2-(trifluoromethyl)phenyl]methyl]pyrazolo[3,4-d]pyrimidine

A mixture of4,6-dichloro-1-(2-(trifluoromethyl)benzyl)-1H-pyrazolo[3,4-d]pyrimidine(50 mg, 144 μmol), 3,3-difluoropyrrolidine hydrochloride (32 mg, 223μmol) and DIPEA (74 mg, 100 μL, 573 μmol) in dioxane (2 mL) was heatedin a microwave oven to 120° C. for 30 min. The solvent was removed underreduce pressure and the crude material was purified by chromatography(silica gel, 20 g, AcOEt/heptane 1/3). Recrystallization from EtOAc andheptane afforded the title compound (35 mg, 58%) as white solid; LC-MS(UV peak area; ESI) 99%, 418.0865 [MH⁺].

Example 384-(3,3-Difluoropyrrolidin-1-yl)-6-(2,2-dimethylpropoxy)-1-[[2-(trifluoromethyl)phenyl]methyl]pyrazolo[3,4-d]pyrimidine

Sodium hydride (6.61 mg, 165 μmol) and 2,2-dimethyl-1-propanol (48.5 mg,59.3 μL, 551 μmol) were added to a solution of6-chloro-4-(3,3-difluoropyrrolidin-1-yl)-1-(2-(trifluoromethyl)benzyl)-1H-pyrazolo[3,4-d]pyrimidine(46 mg, 110 μmol; Example 37 c) in THF (1 mL). The reaction mixture wasstirred for 4 h at 80° C., poured into 25 mL EtOAc and washed with water(2×20 mL). The combined aqueous layers were back-extracted with EtOAc(1×25 mL). The combined organic layers were dried over MgSO₄ andconcentrated in vacuo. The crude material was purified by flashchromatography (silica gel, 20 g, 0% to 50% EtOAc in heptane) to givethe title compound (35 mg, 68%) as white solid; MS (ESI) 470.7 [MH⁺].

Example 39 Pharmacological Tests

The following tests were carried out in order to determine the activityof the compounds of formula (I):

Radioligand Binding Assay

The affinity of the compounds of the invention for cannabinoid CB1receptors was determined using recommended amounts of membranepreparations (PerkinElmer) of human embryonic kidney (HEK) cellsexpressing the human CNR1 or CNR2 receptors in conjunction with 1.5 or2.6 nM [3H]-CP-55,940 (Perkin Elmer) as radioligand, respectively.Binding was performed in binding buffer (50 mM Tris, 5 mM MgCl2, 2.5 mMEDTA, and 0.5% (wt/vol) fatty acid free BSA, pH 7.4 for CB1 receptor and50 mM Tris, 5 mM MgCl₂, 2.5 mM EGTA, and 0.1% (wt/vol) fatty acid freeBSA, pH 7.4 for CB2 receptor) in a total volume of 0.2 ml for 1 h at 30°C. shaking. The reaction was terminated by rapid filtration throughmicrofiltration plates coated with 0.5% polyethylenimine (UniFilter GF/Bfilter plate; Packard). Bound radioactivity was analyzed for Ki usingnonlinear regression analysis (Activity Base, ID Business Solution,Limited), with the Kd values for [3H]CP55,940 determined from saturationexperiments. The compounds of formula (I) show an excellent affinity forthe CB2 receptor with affinities below 10 μM, more particularly of 1 nMto 3 μM and most particularly of 1 nM to 100 nM.

cAMP Assay

CHO cells expressing human CB1 or CB2 receptors are seeded 17-24 hoursprior to the experiment 50.000 cells per well in a black 96 well platewith flat clear bottom (Corning Costar #3904) in DMEM (Invitrogen No.31331), lx HT supplement, with 10% fetal calf serum and incubated at 5%CO₂ and 37° C. in a humidified incubator. The growth medium wasexchanged with Krebs Ringer Bicarbonate buffer with 1 mM IBMX andincubated at 30° C. for 30 min. Compounds were added to a final assayvolume of 100 μl and incubated for 30 min at 30° C. Using thecAMP-Nano-TRF detection kit the assay (Roche Diagnostics) was stopped bythe addition of 50 μl lysis reagent (Tris, NaCl, 1.5% Triton X100, 2.5%NP40, 10% NaN₃) and 50 μl detection solutions (20 μM mAb Alexa700-cAMP1:1, and 48 μM Ruthenium-2-AHA-cAMP) and shaken for 2 h at roomtemperature. The time-resolved energy transfer is measured by a TRFreader (Evotec Technologies GmbH), equipped with a ND:YAG laser asexcitation source. The plate is measured twice with the excitation at355 nm and at the emission with a delay of 100 ns and a gate of 100 ns,total exposure time 10 s at 730 (bandwidth 30 nm) or 645 nm (bandwidth75 nm), respectively. The FRET signal is calculated as follows:FRET=T730-Alexa730-P(T645-B645) with P=Ru730-B730/Ru645-B645, where T730is the test well measured at 730 nM, T645 is the test well measured at645 nm, B730 and B645 are the buffer controls at 730 nm and 645 nm,respectively. cAMP content is determined from the function of a standardcurve spanning from 10 μM to 0.13 nM cAMP.

EC₅₀ values were determined using Activity Base analysis (ID BusinessSolution, Limited). The EC₅₀ values for a wide range of cannabinoidagonists generated from this assay were in agreement with the valuespublished in the scientific literature.

The compounds of the invention are CB2 receptor agonists with EC₅₀ below1 μM and selectivity versus CB1 in the corresponding assay of at least10 fold. Particular compound of the invention are CB2 receptor agonistswith EC₅₀ below 0.01 μM and selectivity versus CB1 in the correspondingassay of at least 2000 fold.

For example, the following compounds showed the following human EC₅₀values in the functional cAMP assay described above:

human CB2 EC₅₀ human CB1 EC₅₀ Example [μM] [μM] 1 0.0048 >10 20.0033 >10 3 0.0051 >10 4 0.0042 >10 5 0.0255 >10 6 0.0059 >10 70.0011 >10 8 0.0054 >10 9 0.0005 >10 10 0.0007 0.3368 11 0.0187 >10 120.0016 1.5741 13 0.0196 >10 14 0.0013 >10 15 0.0009 >10 16 0.0006 >10 170.003 >10 18 0.0262 >10 19 0.0014 >10 20 0.0016 >10 21 0.0034 >10 220.0084 >10 23 0.0055 >10 24 0.0175 >10 25 0.0004 >10 26 0.0356 >10 270.0425 >10 28 0.0019 >10 29 0.0019 >10 30 0.0024 >10 31 0.0173 >10 320.0015 >10 33 0.0088 >10 34 0.005 >10 35 0.0133 >10 36 0.0014 >10 370.0172 >10 38 0.0160 >10

Example A

Film coated tablets containing the following ingredients can bemanufactured in a conventional manner:

Ingredients Per tablet Kernel: Compound of formula (I) 10.0 mg  200.0mg  Microcrystalline cellulose 23.5 mg  43.5 mg  Lactose hydrous 60.0mg  70.0 mg  Povidone K30 12.5 mg  15.0 mg  Sodium starch glycolate 12.5mg  17.0 mg  Magnesium stearate 1.5 mg 4.5 mg (Kernel Weight) 120.0 mg 350.0 mg  Film Coat: Hydroxypropyl methyl cellulose 3.5 mg 7.0 mgPolyethylene glycol 6000 0.8 mg 1.6 mg Talc 1.5 mg 2.6 mg Iron oxide(yellow) 0.8 mg 1.6 mg Titan dioxide 0.8 mg 1.6 mg

The active ingredient is sieved and mixed with microcrystallinecellulose and the mixture is granulated with a solution ofpolyvinylpyrrolidone in water. The granulate is then mixed with sodiumstarch glycolate and magnesium stearate and compressed to yield kernelsof 120 or 350 mg respectively. The kernels are lacquered with an aq.solution/suspension of the above mentioned film coat.

Example B

Capsules containing the following ingredients can be manufactured in aconventional manner:

Ingredients Per capsule Compound of formula (I) 25.0 mg Lactose 150.0mg  Maize starch 20.0 mg Talc  5.0 mg

The components are sieved and mixed and filled into capsules of size 2.

Example C

Injection solutions can have the following composition:

Compound of formula (I) 3.0 mg Polyethylene glycol 400 150.0 mg Aceticacid q.s. ad pH 5.0 Water for injection solutions ad 1.0 ml

The active ingredient is dissolved in a mixture of Polyethylene glycol400 and water for injection (part). The pH is adjusted to 5.0 byaddition of acetic acid. The volume is adjusted to 1.0 ml by addition ofthe residual amount of water. The solution is filtered, filled intovials using an appropriate coverage and sterilized.

The invention claimed is:
 1. A compound of formula (I)

wherein A¹ is CH or N; A² is CH or N; A³ is —(CH₂)_(n)— or —CH₂C(O)—; R¹is alkyl, cycloalkyl, alkoxy or halogen; R² is substituted pyrrolidinylor substituted dihydropyrrolyl, wherein substituted pyrrolidinyl andsubstituted dihydropyrrolyl are pyrrolidinyl and dihydropyrrolylsubstituted with one or two substituents independently selected fromhalogen, hydroxyl, hydroxyalkyl, alkoxyalkyl and alkylfurazanylalkoxy;R³ is phenyl, substituted phenyl, substituted furazanyl, pyridinyl,substituted pyridinyl, dioxothietanyl, tetrahydrofuranyl, substitutedtetrazolyl or substituted triazolyl, wherein substituted phenyl,substituted furazanyl, substituted pyridinyl and substituted triazolylare phenyl, pyridinyl and triazolyl substituted with one or twosubstituents independently selected from alkyl, alkoxy, halogen,haloalkyl, alkylsulfonyl and cycloalkyl, and wherein substitutedtetrazolyl and substituted furazanyl are tetrazolyl and furazanylsubstituted with one substituent selected from alkyl, alkoxy, halogen,haloalkyl, alkylsulfonyl and cycloalkyl; n is 0, 1 or 2; provided thatA¹ and A² are not both CH at the same time; or a pharmaceuticallyacceptable salt or ester thereof.
 2. A compound according to claim 1,wherein A¹ is N.
 3. A compound according to claim 1, wherein A² is N. 4.A compound according to claim 1, wherein A³ is —(CH₂)_(n)—.
 5. Acompound according to claim 1, wherein R¹ is alkyl.
 6. A compoundaccording to claim 1, wherein R¹ is tert.-butyl.
 7. A compound accordingto claim 1, wherein R² is substituted pyrrolidinyl, wherein substitutedpyrrolidinyl is pyrrolidinyl substituted with one or two substituentsindependently selected from halogen and hydroxyl.
 8. A compoundaccording to claim 1, wherein R² is difluoropyrrolidinyl orhydroxypyrrolidinyl.
 9. A compound according to claim 1, wherein R³ issubstituted phenyl, substituted furazanyl, substituted pyridinyl,substituted tetrazolyl or substituted triazolyl, wherein substitutedphenyl, substituted pyridinyl and substituted triazolyl are phenyl,pyridinyl and triazolyl substituted with one or two substituentsindependently selected from alkyl, halogen and haloalkyl, whereinsubstituted tetrazolyl is tetrazolyl substituted with one substituentselected from alkyl, and cycloalkyl, and wherein substituted furazanylis furazanyl substituted with alkyl.
 10. A compound according to claim1, wherein R³ is trifluoromethylphenyl, methylfurazanyl,chloropyridinyl, methyltetrazolyl, cyclopropyltetrazolyl,dimethyltriazolyl or methyltriazolyl.
 11. A compound according to claim1, wherein n is
 1. 12. A compound selected from the group consisting of:6-tert-Butyl-4-(3,3-difluoro-pyrrolidin-l-yl)-[(4-methoxyphenyl)methyl]-1H-pyrazolo[3,4-d]pyrimidine;1-Benzyl-6-tert-butyl-4-(3,3-difluoro-pyrrolidin-1-yl)-1H-pyrazolo[3,4-d]pyrimidine;(S)-1-[6-tert-Butyl-1-[(2-chlorophenyl)methyl]-1H-pyrazolo[3,4-d]pyrimidin-4-yl]-pyrrolidin-3-ol;6-tert-Butyl-1-[(2-chlorophenyl)methyl]-4-(3,3-difluoro-pyrrolidin-1-yl)-1H-pyrazolo[3,4-d]pyrimidine;6-tert-Butyl-1-[(2-chlorophenyl)methyl]-4-ethoxy-1H-pyrazolo[3,4-d]pyrimidine;6-tert-Butyl- 1-[(2-chloro-4-fluorophenyl)methyl ]-4-(3,3-difluoro-pyrrolidin-1-yl)-1H-pyrazolo[3,4-d]pyrimidine;6-tert-Butyl-4-(3,3-difluoro-pyrrolidin-1-yl)-1-[(2-trifluoromethylphenyl)methyl]-1H-pyrazolo[3,4-d]pyrimidine;6-tert-Butyl-4-(3,3-difluoro-pyrrolidin-1-yl)-1-phenethyl-1H-pyrazolo[3,4-d]pyrimidine;6-tert-Butyl-4-(3,3-difluoro-pyrrolidin-1-yl)-1-(4-methyl-furazan-3-ylmethyl)-1H-pyrazolo[3,4-d]pyrimidine;6-tert-Butyl-4-(3,3-difluoro-pyrrolidin-1-yl)-1-(2-methanesulfonyl-benzyl)-1H-pyrazolo[3,4-d]pyrimidine;6-tert-Butyl-4-(3,3-difluoro-pyrrolidin-1-yl)-1-(2-pyridin-3-yl-ethyl)-1H-pyrazolo[3,4-d]pyrimidine;6-tert-Butyl-1-(2-chloro-pyridin-3-ylmethyl)-4-(3,3-difluoro-pyrrolidin-1-yl)-1H-pyrazolo[3,4-d]pyrimidine; 6-tert-Butyl-4-(3,3-difluoro-pyrrolidin- 1-yl)-1-(1,1-dioxo-1λ6-thietan-3-yl)-1H-pyrazolo [3,4-d]pyrimidine;6-tert-Butyl-4-(3,3-difluoro-pyrrolidin-1-yl)-1-(tetrahydro-furan-3-yl)-1H-pyrazolo[3,4-d]pyrimidine; 6-tert-Butyl-4-(3,3-difluoro-pyrrolidin-1-yl)-1-(1-methyl- 1H-tetrazol-5-ylmethyl)-1H-pyrazolo[3,4-d]pyrimidine;6-tert-Butyl -1-(1-cyclopropyl-1H-tetrazol-5-ylmethyl)-4-(3,3-difluoro-pyrrolidin-1-yl)-1H-pyrazolo[3,4-d]pyrimidine;6-tert-Butyl-4-(3,3-difluoro-pyrrolidin- 1-yl)-1-(4,5-dimethyl-4H-[1,2,4]triazol-3-ylmethyl)-1H-pyrazolo[3,4-d]pyrimidine;6-tert-Butyl-1-(4,5-dimethyl-4H-[1,2,4]triazol-3-ylmethyl)-4-(3-fluoro-2,5-dihydro-pyrrol-1-yl)-1H-pyrazolo[3,4-d]pyrimidine;6-tert-Butyl-4-(3,3-difluoro-pyrrolidin- 1-yl)- 1-(3-methyl-3H-[1,2,3]triazol-4-ylmethyl)-1H-pyrazolo[3,4-d]pyrimidine;6-tert-Butyl-4-(3-fluoro-2,5-dihydro-pyrrol- 1-yl)-1-(3-methyl-3H-[1,2,3 ]triazol-4-ylmethyl)-1H-pyrazolo[3,4-d]pyrimidine;2-[6-tert-Butyl-4-(3,3-difluoro-pyrrolidin-1-yl)-pyrazolo[3,4-d]pyrimidin-1-yl]-1-pyridine-4-yl-ethanone;2-[6-tert-Butyl-4-(3,3-difluoro-pyrrolidin-1-yl)-pyrazolo[3,4-d]pyrimidin-1-yl]-1-pyridine-2-yl-ethanone;(S)-1-[6-tert-Butyl-1-[(4-methoxyphenyl)methyl]-1H-pyrazolo[3,4-d]pyrimidin-4-yl]-pyrrolidin-3-ol;6-Cyclopropyl-4-(3,3-difluoro-pyrrolidin-1-yl)-1-[(4-methoxyphenyl)methyl]-1H-pyrazolo [3,4-b]pyridine; (S)-1-[6-tert-Butyl-1-(4-methyl-furazan-3-ylmethyl)-1H-pyrazolo[3,4-d]pyrimidin-4-yl]-pyrrolidin-3-ol;6-tert-Butyl-4-[(S)-3-(4-methyl-furazan-3-ylmethoxy)-pyrrolidin-1-yl]-1-(4-methyl-furazan-3-ylmethyl)- 1H-pyrazolo[3,4-d]pyrimidine;6-Cyclopropyl-4-(3,3-difluoro-pyrrolidin- 1-yl)-1-(4-methyl-furazan-3-ylmethyl)-1H-pyrazolo[4,3-c]pyridine;6-Cyclopropyl-4-(3,3-difluoro-pyrrolidin-1-yl)-1-(4-methyl-furazan-3-ylmethyl)-1H-pyrazolo[3,4-b]pyridine;(S)-1-[6-tert-Butyl -1-[(2-trifluoromethylphenyl)methyl]-1H-pyrazolo[3,4-d]pyrimidin-4-yl]-pyrrolidin-3-ol;(S)-1-[6-tert-Butyl-1-(2-methanesulfonyl-benzyl)- 1H-pyrazolo[3,4-d]pyrimidin-4-yl]-pyrrolidin-3-ol; (S)-1-[6-tert-Butyl-1-(1-methyl- 1H-tetrazol-5-ylmethyl)-1H-pyrazolo[3,4-d]pyrimidin-4-yl]-pyrrolidin-3-ol;(S)-1-[6-tert-Butyl-1-(3-chloro-pyridin-2-ylmethyl)-1H-pyrazolo[3,4-d]pyrimidin-4-yl]-pyrrolidin-3-ol;(S)-1-[6-tert-Butyl-1-(1-cyclopropyl-1H-tetrazol-5-ylmethyl)-1H-pyrazolo[3,4-d]pyrimidin-4-yl]-pyrrolidin-3-ol;{(R)-1-[6-tert-Butyl-1-[(2-trifluoromethylphenyl)methyl]-1H-pyrazolo[3,4-d]pyrimidin-4-yl]-pyrrolidin-2-yl}-methanol;6-tert-Butyl-4-((R)-2-methoxymethyl-pyrrolidin- 1-yl)-1-[(2-trifluoromethylphenyl)methyl]-1H-pyrazolo[3,4-d]pyrimidine;6-tert-Butyl-4-((R)-2-methoxymethyl-pyrrolidin- 1-yl)-1-(4-methyl-furazan-3-ylmethyl)-1H-pyrazolo[3,4-d]pyrimidine;6-Chloro-4-(3,3-difluoropyrrolidin-1-yl)-1-[[2-(trifluoromethyl)phenyl]methyl]pyrazolo[3,4-d]pyrimidine; and 4-(3,3-Difluoropyrrolidin- 1-yl)-6-(2,2-dimethylpropoxy)- 1-[[2-(trifluoromethyl)phenyl]methyl]pyrazolo[3,4-d]pyrimidine.
 13. Acompound according to claim 1 selected from the group consisting of:6-tert-Butyl-4-(3,3-difluoro-pyrrolidin-1-yl)-1-[(2-trifluoromethylphenyl)methyl]-1H-pyrazolo[3,4-d]pyrimidine;6-tert-Butyl-4-(3,3-difluoro-pyrrolidin-1-yl)-1-(4-methyl-furazan-3-ylmethyl)- 1H-pyrazolo[3,4-d]pyrimidine;6-tert-Butyl-1-(2-chloro-pyridin-3-ylmethyl)-4-(3,3-difluoro-pyrrolidin-1-yl)-1H-pyrazolo[3,4-d]pyrimidine;6-tert-Butyl-4-(3,3-difluoro-pyrrolidin-1-yl)-1-(1-methyl-1H-tetrazol-5-ylmethyl)-1H-pyrazolo[3,4-d]pyrimidine;6-tert-Butyl-1-(1-cyclopropyl-1H-tetrazol-5-ylmethyl)-4-(3,3-difluoro-pyrrolidin-1-yl)-1H-pyrazolo[3,4-d]pyrimidine;6-tert-Butyl-4-(3,3-difluoro-pyrrolidin-1-yl)-1-(4,5-dimethyl-4H-[1,2,4]triazol-3-ylmethyl)-1H-pyrazolo[3,4-d]pyrimidine;6-tert-Butyl-4-(3,3-difluoro-pyrrolidin-1-yl)-1-(3-methyl-3H-[1,2,3]triazol-4-ylmethyl)-1H-pyrazolo[3,4-d]pyrimidine;(S)-1-[6-tert-Butyl-1-(4-methyl-furazan-3-ylmethyl)-1H-pyrazolo[3,4-d]pyrimidin-4-yl]-pyrrolidin-3-ol;and(S)-1-[6-tert-Butyl-1-[(2-trifluoromethylphenyl)methyl]-1H-pyrazolo[3,4-d]pyrimidin-4-yl]-pyrrolidin-3-ol.14. A process for the preparation of a compound according to claim 1,comprising one of the following steps: (a) the reaction of a compound offormula (A)

in the presence of R²H and a base; (b) the reaction of a compound offormula (B)

in the presence of R³-A³-X and a base; (c) the reaction of a compound offormula (C)

in the presence of MR¹, a palladium catalyst and a base; or (d) thereaction of a compound of formula (D)

in the presence of MR¹, a palladium catalyst and a base; wherein A¹ toA³ and R¹ to R³ are as defined in claim 1, X is a leaving group and M isa suitably substituted metal species.
 15. A compound manufacturedaccording to a process of claim
 14. 16. A pharmaceutical compositioncomprising a compound in accordance with claim 1 and a therapeuticallyinert carrier.